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Development and Evaluation of an Antisense Acc-Oxidase (CMACO-a) Galia F1 Hybrid Muskmelon (Cucumis melo L. Var. Reticul...

Permanent Link: http://ufdc.ufl.edu/UFE0023789/00001

Material Information

Title: Development and Evaluation of an Antisense Acc-Oxidase (CMACO-a) Galia F1 Hybrid Muskmelon (Cucumis melo L. Var. Reticulatus Ser.)
Physical Description: 1 online resource (210 p.)
Language: english
Creator: Harty, Jeanmarie
Publisher: University of Florida
Place of Publication: Gainesville, Fla.
Publication Date: 2009

Subjects

Subjects / Keywords: agriculture, aroma, postharvest, protected, sensory, volatiles
Horticultural Science -- Dissertations, Academic -- UF
Genre: Horticultural Science thesis, Ph.D.
bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: Recognized for its flavor, the Galia muskmelon (Cucumis melo L. var. reticulatus Ser.) is also known to have a short shelf-life. To address this concern, previous work transformed the Galia male parental line with an antisense ACC oxidase (CMACO-1) gene. The gene inhibited the last step in ethylene biosynthesis, resulting in transgenic male parental lines that produced less ethylene and were firmer, subsequently possessing a longer shelf-life than the non-transgenic fruit. These lines were used to create antisense Galia (ASG) hybrids (ASxWT, ASxAS, WTxAS). During fall 2006, preliminary ASG lines were evaluated with the original Galia muskmelon. Fruits were harvested at four stages: stage 1.) zero-slip, green (ZG); 2.) zero-slip, yellow-green (ZYG); 3.) half-slip (HS); and 4.) full-slip (FS). Data were recorded for days to harvest, fruit size, quality, ethylene evolution, and respiration. At stage ZG, all ASG muskmelons were similar in size, quality, ethylene and respiration to Galia . At stage ZYG, ASxAS and ASxWT muskmelons were significantly firmer than Galia and produced less ethylene yet were similar in soluble solids content (SSC). On average, ASG melons remained on the vine six days longer than Galia and were similar in quality to Galia at stages HS and FS. Aroma volatiles were identified in order to determine what sets Galia flavor apart from look-a-like cultivars. The compounds considered important in Galia muskmelons were benzyl acetate, ethyl-2-methyl butyrate, methyl 2-methyl butyrate, ethyl isobutyrate, 2-methylbutyl acetate, hexyl acetate, ethyl butyrate, ethyl caproate, cis-3-hexenyl acetate, and isovaleronitrile. ASG and Galia muskmelon aroma was evaluated in 2006 and 2007. The greatest differences in aroma among ASG and Galia were at stage ZYG, where volatiles were greatest in Galia . After five-days storage at 20 C in fall 2007, line ASxAS fruit remained firmer, when harvested at stage ZYG. At stages ZG, HS and FS, aroma and quality differences were few. On average, ASG fruit remained on the vine four days longer than Galia , suggesting a wider harvest window. Even though there were some differences in volatiles at stage ZYG, in order to enhance shipping quality, it is recommended that line ASxAS be harvested at stage ZYG, where SSC was acceptable and fruit firmness was greatest at harvest and after storage.
General Note: In the series University of Florida Digital Collections.
General Note: Includes vita.
Bibliography: Includes bibliographical references.
Source of Description: Description based on online resource; title from PDF title page.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Statement of Responsibility: by Jeanmarie Harty.
Thesis: Thesis (Ph.D.)--University of Florida, 2009.
Local: Adviser: Cantliffe, Daniel J.
Electronic Access: RESTRICTED TO UF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE UNTIL 2011-05-31

Record Information

Source Institution: UFRGP
Rights Management: Applicable rights reserved.
Classification: lcc - LD1780 2009
System ID: UFE0023789:00001

Permanent Link: http://ufdc.ufl.edu/UFE0023789/00001

Material Information

Title: Development and Evaluation of an Antisense Acc-Oxidase (CMACO-a) Galia F1 Hybrid Muskmelon (Cucumis melo L. Var. Reticulatus Ser.)
Physical Description: 1 online resource (210 p.)
Language: english
Creator: Harty, Jeanmarie
Publisher: University of Florida
Place of Publication: Gainesville, Fla.
Publication Date: 2009

Subjects

Subjects / Keywords: agriculture, aroma, postharvest, protected, sensory, volatiles
Horticultural Science -- Dissertations, Academic -- UF
Genre: Horticultural Science thesis, Ph.D.
bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: Recognized for its flavor, the Galia muskmelon (Cucumis melo L. var. reticulatus Ser.) is also known to have a short shelf-life. To address this concern, previous work transformed the Galia male parental line with an antisense ACC oxidase (CMACO-1) gene. The gene inhibited the last step in ethylene biosynthesis, resulting in transgenic male parental lines that produced less ethylene and were firmer, subsequently possessing a longer shelf-life than the non-transgenic fruit. These lines were used to create antisense Galia (ASG) hybrids (ASxWT, ASxAS, WTxAS). During fall 2006, preliminary ASG lines were evaluated with the original Galia muskmelon. Fruits were harvested at four stages: stage 1.) zero-slip, green (ZG); 2.) zero-slip, yellow-green (ZYG); 3.) half-slip (HS); and 4.) full-slip (FS). Data were recorded for days to harvest, fruit size, quality, ethylene evolution, and respiration. At stage ZG, all ASG muskmelons were similar in size, quality, ethylene and respiration to Galia . At stage ZYG, ASxAS and ASxWT muskmelons were significantly firmer than Galia and produced less ethylene yet were similar in soluble solids content (SSC). On average, ASG melons remained on the vine six days longer than Galia and were similar in quality to Galia at stages HS and FS. Aroma volatiles were identified in order to determine what sets Galia flavor apart from look-a-like cultivars. The compounds considered important in Galia muskmelons were benzyl acetate, ethyl-2-methyl butyrate, methyl 2-methyl butyrate, ethyl isobutyrate, 2-methylbutyl acetate, hexyl acetate, ethyl butyrate, ethyl caproate, cis-3-hexenyl acetate, and isovaleronitrile. ASG and Galia muskmelon aroma was evaluated in 2006 and 2007. The greatest differences in aroma among ASG and Galia were at stage ZYG, where volatiles were greatest in Galia . After five-days storage at 20 C in fall 2007, line ASxAS fruit remained firmer, when harvested at stage ZYG. At stages ZG, HS and FS, aroma and quality differences were few. On average, ASG fruit remained on the vine four days longer than Galia , suggesting a wider harvest window. Even though there were some differences in volatiles at stage ZYG, in order to enhance shipping quality, it is recommended that line ASxAS be harvested at stage ZYG, where SSC was acceptable and fruit firmness was greatest at harvest and after storage.
General Note: In the series University of Florida Digital Collections.
General Note: Includes vita.
Bibliography: Includes bibliographical references.
Source of Description: Description based on online resource; title from PDF title page.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Statement of Responsibility: by Jeanmarie Harty.
Thesis: Thesis (Ph.D.)--University of Florida, 2009.
Local: Adviser: Cantliffe, Daniel J.
Electronic Access: RESTRICTED TO UF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE UNTIL 2011-05-31

Record Information

Source Institution: UFRGP
Rights Management: Applicable rights reserved.
Classification: lcc - LD1780 2009
System ID: UFE0023789:00001


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1 DEVELOPMENT AND EVALUATION OF AN ANTISENSE ACC OXIDASE (CMACO 1) GALIA F1 HYBRID MUSKMELON (Cucumis melo L. var. reticulatus Ser .) By JEANMARIE MINK HARTY A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSI TY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2009

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2 2009 Jeanmarie Mink Harty

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3 To my husband, Cheyenne, for tireless support, advice and so much love. A nd, to my parents, who have always supported me and have been instrumental in my pursuit of a career in agriculture. Without you, I would not be where I am. I love and thank you.

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4 ACKNOWLEDGMENTS This dissertation could not have been accomplished without the help of many people. I am most grateful to my committee chair and advisor, Daniel J. Cantliffe for his wisdom, guidance, support, expertise and patience throughout my graduate work. I am also appreciative of my graduate comm ittee members, Dr. Harry J. Klee for expertise in DNA, plant breeding and aroma; Dr. Steven A. Sargent for providing postharvest guidance; Dr. Peter J. Stoffella for his proficient knowledge in statistics and project design; and Dr. Lawrence Datnoff fo r education in plant pathology, data collection and analysis. Each one of my committee members have been kind, patient and enabled me to gain a great deal of knowledge from them. None of this research could have been done without the direction of Nicole S haw Pratt, who taught me how to grow Galia muskmelons and was always there to assist me when I asked for help. Special thanks must go to Denise Tieman, whose patience and tolerance helped me learn how to collect aroma volatiles, integrate peaks and unde rstand how a GC works. And without the assistance of Melissa Webb, I would not have made many important deadlines. I t hank the many past and present members of the Building 710 Lab that I had the opportunity and pleasure to work with throughout the yea rs. These people included: Dr. Elio Jovicich, Dr. Hector Nunez -Paleius, Dr. Silvia Rondon, Dr. Ivanka Kozareva, Elizabeth Thomas, Jiyoung Hong, Dr. Dzingai Rukuni, Dr. Shubin Saha, Jimmy Webb and Jennifer Noseworthy I would like to express my gr atitud e to the postharvest crew ( including Dr. Donald Huber, Kim Cordasco, Adrian Berry, Dr. Brandon Hurr, Eunkyung Lee, Sharon Dea, Marcio Eduardo Canto Pereira Oren Warren and Sherry (Ming Wei) Kao ) for always

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5 allowing me into their labs and full use of equipment (sometimes as early as three a.m. ). I thank the members of the Settles lab (including Drs. Mark Settles, Chiwah Seung and Diego Fajardo ) for allowing me to extract DNA and introducing me to new extraction technology. I thank the Klee Lab ( including Peter Bliss and Drs. M ark Taylor, Brian Kevani, Michelle Zeigler Val Dal Cin Sandrine and Jonathan Vogel ) who were always friendly and accommodating when I had any question, required assistance or nee ded to run a PCR. Also, thank you to Dr.Charlie Sims and his assistant, Lorenzo Puentes, for the melon sensory panel. A very special thanks goes to Gene Hannah, whose friendship and assistance at the greenhouse was beyond measure and much appreciated. I also thank John Thomas and Cecil Shine for great advi ce help and support. And, a heartfelt thanks go to the entire Horticultural Sciences faculty, staff and graduate students whose smiles, kind words and assistance were always nearby. Finally, I would like to recognize the Tropical/Sub T ropical Agricult ural Research (TSTAR) grant program for funding this research and the University of Florida Graduate School, Office of Graduate and Minority Programs for the Graduate Supplemental Tuition Scholarship, which provided my tuition in summer and fall, 2008 and spring, 2009.

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6 TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................................................................................................... 4 LIST OF TABLES ............................................................................................................................... 9 LIST OF FIGURES ............................................................................................................................ 12 ABSTRACT ........................................................................................................................................ 14 CHAPTER 1 INTRODUCTION ...................................................................................................................... 16 2 LITERATURE REVIEW ........................................................................................................... 20 Introduction ................................................................................................................................. 20 Classification and Origin .................................................................................................... 20 Consumption and Produc tion ............................................................................................. 24 Characteristics ............................................................................................................................. 25 Climacteric and Non Climacteric Fruit .............................................................................. 26 Maturity and Ripening ........................................................................................................ 29 Fruit Quality ................................................................................................................................ 30 Sweetness ............................................................................................................................. 31 Acids ..................................................................................................................................... 32 Texture ................................................................................................................................. 32 Color ..................................................................................................................................... 33 Aroma ................................................................................................................................... 34 Taste and Sensory Analysis ................................................................................................ 40 Galia Muskmelon ..................................................................................................................... 41 Galia and Galia Type Specialty Cultivars .................................................................... 41 Galia Muskmelon Production .......................................................................................... 42 Galia Muskmelon Postharvest Practices ......................................................................... 45 3 PRODUCTION, EVALUATION, AND SELECTION OF ANTISENSE ACC OXIDASE (CMACO 1) GALIA F1 HYBRID MUSKMELON ( Cucumis melo L. var. reticulatus Ser.) ............................................................................................................... 49 Introduction ................................................................................................................................. 49 Materials and Methods ............................................................................................................... 52 Antisense ACC -oxidase Galia F1 Hybrid Development ................................................ 52 T ransgene Detection for Antisense Male, Female and Hybrid Lines .............................. 54 ASG Muskmelon Production .............................................................................................. 54 Fruit Harvest Procedure ...................................................................................................... 55 Ethylene, Respiration and Fruit Quality Measurements ................................................... 56 Statistical Analysis .............................................................................................................. 56

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7 Results and Discussion ............................................................................................................... 57 Parental Line Production ..................................................................................................... 57 Hybrid Muskmelon Results and Selection, Fall 2006 ....................................................... 58 Fall 2006 and Spring 2007 ASG 1 Results ........................................................................ 62 Stage ZG ....................................................................................................................... 62 Stage ZYG .................................................................................................................... 62 Stage HS ....................................................................................................................... 63 Stage FS ........................................................................................................................ 63 Summary ...................................................................................................................................... 65 4 GALIA MUSKMELON FRUIT QUALITY AND FLAVOR ( Cucumis melo L. var. reticulatus Ser.) .................................................................................................................... 80 Introduction ................................................................................................................................. 80 Materials and Methods ............................................................................................................... 83 Fruit Selection and Postharvest Treatments ....................................................................... 86 Ethylene and Respiration Measurements ........................................................................... 87 Fruit Quality Measurements ............................................................................................... 87 Aroma Volatile Collection and Analysis ........................................................................... 88 Sensory Evaluation .............................................................................................................. 89 Statistical Analysis .............................................................................................................. 91 Results .......................................................................................................................................... 91 Days to Harves t (DTH) ................................................................................................ 91 Fruit Quality and Aroma Volatiles ............................................................................. 92 Stage ZG ....................................................................................................................... 92 Stag e ZYG .................................................................................................................... 94 Stages HS ...................................................................................................................... 96 Stage FS ........................................................................................................................ 97 Sensory analysis .................................................................................................................. 99 Summary .................................................................................................................................... 1 04 5 AROMA VOLATILE AND FRUIT QUALITY EVALUATION OF ANTISENSE ACC OXIDASE (CMACO 1) GALIA F1 HYBRID MUSKMELONS ( Cucumis melo L. v ar. reticulatus Ser.) ..................................................... 117 Materials and Methods ............................................................................................................. 119 Fruit Selection and Postharvest Treatments ..................................................................... 121 Ethylene and Respiration Measurements ......................................................................... 122 Fruit Quality Measurements ............................................................................................. 122 Statistical Analysis ............................................................................................................ 124 Results and Discussion ............................................................................................................. 124 Days to Harvest ................................................................................................................. 124 Fruit Qualit y and Aroma ................................................................................................... 125 Stage ZG ..................................................................................................................... 125 Stage ZYG .................................................................................................................. 127 Stage HS ..................................................................................................................... 131 Stage FS ...................................................................................................................... 133

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8 Summary .................................................................................................................................... 137 6 CONCLUSIONS ....................................................................................................................... 150 APPENDIX A CHAPTER 3 ANOVA TABLES ............................................................................................. 156 Additional Tables for Chapter 3 ............................................................................................... 160 B ADDITIONAL TABLES AND ANOVA TABLES FOR CHAPTER 4 .............................. 162 Appendix B 1 ............................................................................................................................ 162 Appendix B 2 ............................................................................................................................ 167 Appendix B 3 ............................................................................................................................ 172 Appendix B 4 ............................................................................................................................ 177 Chapter 4 ANOVA Tables ....................................................................................................... 182 C CHAPTER 5 ANOVA TABLES ............................................................................................. 191 LIST OF REFERENCES ................................................................................................................. 195 BIOGRAPHICAL SKETCH ........................................................................................................... 210

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9 LIST OF TABLES Table page 2 1 List of Galia type muskmelon ( Cucumis melo L. var. reticulatus Ser.) cultivars. ................................................................................................................................. 48 3 1 Days to harves t (DTH) results of the transformed (T) antisense male (TGM AS 1 and TGM -AS 2) lines that were selfed from spring 2004 through fall 2005. ....................................................................................................................................... 67 3 2 Days to harvest (DTH) results of backcrossed (BC ) antisense female (TGF AS 1 and TGF -AS 2) lines from spring 2004 through fall 2005. ....................................... 67 3 3 Stage ZG means of days to harvest, fruit quality, ethylene production and respiration rates of Gali a and grouped Antisense Galia (ASG 1 and 2) muskmelons, fall 2006. .......................................................................................................... 68 3 4 Stage ZYG means of days to harvest, fruit quality, ethylene production and respiration rates of Galia, and indiv idual lines of Antisense Galia (ASG -1 and 2) muskmelons, fall 2006. .............................................................................................. 69 3 5 Stage HS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia, Galia ty pe (MG10183) and Antisense Galia (ASG 1 and 2) muskmelons, fall 2006. ............................................................................... 70 3 6 Stage FS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antis ense Galia (ASG 1 and 2) muskmelons, fall 2006. .......................................................................................................... 71 3 7 Stage ZG means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antisense Galia (ASG 1) muskmelons, fall 2006 and spring 2007. ............................................................................................................ 72 3 8 Stage ZYG means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antisense Galia (ASG 1) muskmelons, fall 2006 and spring 2007. ............................................................................................................ 73 3 9 Stage ZYG line x season (L x S) interaction means of soluble solids content, ethylene production and respiration rates. ............................................................................ 73 3 10 Stage HS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antisense Galia (ASG 1) muskmelons, fall 2006 and spring 2007. ............................................................................................................ 74 3 11 Stage HS line x season (L x S) interaction means of fruit length, soluble solids content and ethylene. ................................................................................................... 74

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10 3 12 Stage FS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antisense Galia (ASG 1) muskmelons, fall 2006 and spring 2007. ............................................................................................................ 75 3 13 Stage FS means of significant line*season interaction means of days to harvest. .................................................................................................................................... 76 4 1 Volatile compounds considered to be significant contributors to the aroma of Galia and GT cultivars, MG10183 and Elario, fall 2006, spring 2007 and fall 2007. ............................................................................................................................... 107 4 2 Stage ZG days to harvest and fruit quality means at harvest of Galia and Galia -type muskmelons. ................................................................................................... 108 4 3 Stage ZG means of to tal identified volatiles (TIV), measured in ng gFW1 h1, from Galia and Galia type muskmelons. ...................................................................... 108 4 4 Stage ZYG fruit quality means of Galia and Galia type muskmelons. ...................... 109 4 5 Stage ZYG means of total identified volatiles (TIV), measured in ng gFW1 h1, from Galia and Galia type muskmelons. ................................................................... 109 4 6 Stage HS fruit quality means of Galia and Galia type muskmelons. .......................... 110 4 7 Stage HS means, measured in ng gFW1 h1, of total identified volatiles (TIV), from Galia and Galia type muskmelons. ...................................................................... 110 4 8 Stage FS fruit quality means of Galia and Galia type muskmelons. .......................... 111 4 9 Stage FS means of total identified volatil es (TIV), measured in ng gFW1 h1, from Galia and Galia type muskmelons. ...................................................................... 111 4 10 Temperatures and photosynthetic photon flux ( PPF) during fall 2006, spring and fall 2007 of Galia and Galia type muskmelons grown in a passively ventilated greenhouse. ......................................................................................................... 112 4 11 Stage FS means of soluble solids content (SSC, Brix), firmness (N) and aroma (ng gFW1 h1) of Galia, Galia ty pe and Red Moon melons, spring 2008. .......................................................................................................................... 113 5 1 Odor detection threshold levels (OTV) of 17 the significant contributor aroma compounds of Galia and ASG muskmelons (adapted from Mitchell Hart y et al., 2008). ............................................................................................................... 139 5 2 Stage ZG means of days to harvest and fruit quality variables for Galia and ASG lines at harvest, fall 2006, spring and fall 2007. ....................................................... 140

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11 5 3 Stage ZYG means of days to harvest and fruit quality variables for Galia and ASG lines at harvest, fall 2006, spring and fall 2007. ................................................ 141 5 4 Stage ZYG me ans of significant line*season fruit quality parameters at harvest. .................................................................................................................................. 142 5 5 Stage HS means of days to harvest and fruit quality variables for Galia and ASG lines at harvest, fall 2006, spring and fall 2007. ....................................................... 143 5 6 Stage HS means of significant line*season interaction of TA, pH and ethylene at harvest. ............................................................................................................... 144 5 7 Stage FS means of fruit quality variables for Galia and ASG lines at harvest, fall 2006, spring and fall 2007. ............................................................................. 145 5 8 Stage FS means of significant line*season interaction for days to harvest, we ight and length ................................................................................................................. 146 5 9 Means of total identified volatiles (TIV) (ng g FW1 hr1) at harvest for Galia and ASG muskmelons harvested at stages ZG, ZYG, HS and FS in fall 2006, spring and fal l 2007. ........................................................................................... 147 5 10 Means of total identified volatiles (TIV) (ng g FW1 hr1) after storage at 20 C for Galia and ASG muskmelons harvested at stages ZG, ZYG, HS and FS in spring and fall 2007. .................................................................................................. 147 5 11 Stage ZYG means, presented in ng gFW1 h1 of Galia and Antisense Galia (ASG) muskmelon aroma compounds measured at harvest, fall 2006, spring 2007 and fall 2007. .............................................................................................................. 148

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12 LIST OF FIGURES Figure page 3 1 Galia muskmelon parental lines. The female parental line, Noy Yizreel (Left) and the male parental line, Krymka (righ t). ............................................................ 76 3 2 The Galia F1 hybrid muskmelon. ....................................................................................... 76 3 3 TGM -AS 1 fruit from a T4 generation still green and on the vine after pollina tion on 3 23 06; and a wild-type male fruit ready to be harvested at full -slip that was pollinated on 3 25 06. ............................................................................... 77 3 4 Ethylene evolution and respiration (CO2) rates of T3 TGM -AS 1 muskmelons h arvested at different stages (ZG= zero -slip, green; ZYG= zero -slip, yellow/green; HS= half -slip; FS= full -slip; PS= post -slip) of ripening, fall 2005. ....................................................................................................................................... 77 3 5 Ethylene evolution and (CO2) respi ration rates of T3 TGM -AS 2 muskmelons harvested at different stages (ZG= zero -slip, green; ZYG= zero -slip, yellow/green; HS= half -slip; FS= full -slip; PS= post -slip) of ripening, fall 2005. ....................................................................................................................................... 78 3 6 Ethylene evolution and respiration (CO2) rates of wild type male (Krymka) muskmelons harvested at different stages (ZG= zero -slip, green; ZYG= zero slip, yellow/green; HS= half -slip; FS= full -slip; PS= post -slip) of ripening, fall 2005. ................................................................................................................................. 78 3 7 The Protected Agriculture Greenhouse site enveloped in smoke from nearby wildfires during the week of May 8, 2007. .......................................................................... 79 4 1 Ethylene a nd respiration rates during storage at 20 C harvested at stages ZG, ZYG, HS and FS for Galia and GT muskmelons, fall 2007. .................................. 114 4 2 Aroma volatile emissions of 17 SC compounds found in Ga lia, MG10183 and Elario, fall 2007, (Number (n) of fruits ranged from 1 to 12). 1= benzaldehyde, 2= isovaleronitrile, 3= isobutyl propionate, 4= ethyl 3 (methylthio)propionate, 5= amyl acetate, 6= cis 6 -nonen1 ol, 7= ethyl caproate, 8= benzyl acetate 9= ethyl propionate, 10= ethyl isobutyrate, 11= isobutyl acetate, 12 = propyl acetate, 13= hexyl acetate, 14= ethyl butyrate, 15= butyl acetate, 16= ethyl 2 -methyl butyrate, 17= 2-methylbutyl acetate. .................. 115 4 3 Galia, MG10183, Elario and Red Moon melons ( Cucumis melo L.)used in the sensory panel, spring 2008. ......................................................................... 116 4 4 Sensory evaluation results from fruit harvested at the recommended stage, FS for Galia, MG10183 and Elario; and the recommended harvest stage for

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13 Red Moon, which is when a crack begins at the abscission layer, spring 2008. ..................................................................................................................................... 116 5 1 Average, maximum and minimum temperatures and solar radiation (Photsynthetic Photon Flux (PPF)) for Galia and antisense Galia (ASG) produced in a passively -ventilated greenhouse, fall 2006, spring and fall 2007. ..................................................................................................................................... 149

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14 Abstract of Dissertation Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy DEVELOPMENT AND EVALUATION OF AN ANTISENSE ACC OXIDASE (CMACO 1) GALIA F1 HYBRID MUSKMELON (Cucumis melo L. var. reticulatus Ser .) By Jeanmarie Mink Harty May 2009 Chair: Daniel J. Cantliffe Major: Horticultural Science Recognized for its flavor, the Galia muskmelon ( Cucumis melo L. var. r eticulatus Ser.) is also know n to have a short shelf -life. To address this concern, p r evious work transformed the Galia male parental line with an antisense ACC oxidase ( CM ACO 1) gene The gene inhib it ed the last step in ethylene biosynthesis resulting in transgenic male parental lines that produced less ethylene and were firmer subsequently possessing a longer shelf -life than the non trans genic fruit These lines were used to create antisense Galia (ASG) hybrid s (ASxWT ASxAS WT xAS). During fall 2006, preliminary ASG lines were evalua ted with the orig inal Galia muskmelon Fruits were harv ested at four stages : s tage 1.) zero -slip, green (ZG); 2.) zero slip, yellow -green (ZYG); 3.) half -slip (HS); and 4.) full -slip (FS). Data were recorded for days to harvest, fruit size, quality, ethy lene evolution and respiration At stage ZG, all ASG musk melons were similar in size, quality, ethylene and respiration to Galia At stage ZYG, ASxAS and ASxWT musk melons were s ignificantly firmer than Galia and produced less ethylene yet were similar in soluble solids content (SSC). On average,

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15 ASG m elons remained on the vine six days longer than Galia and were similar in quality to Galia at stages HS and FS Aroma volatiles were identified in order to determine what sets Galia flavor apart from look a -like cultivars. T he compounds considered important in Galia muskmelons were benzyl acetate, ethyl 2 -methyl butyrat e, methyl 2 -methyl butyrate, ethyl isobutyrate 2 -methylbutyl acetate hexyl acetate, ethyl butyrate ethyl caproate, cis 3 hexenyl acetate and isovaleronitrile ASG and Galia muskmelon aroma was evaluated in 2006 and 2007. The greatest differences in aroma among ASG and Galia were at stage ZYG where volatiles were greatest in Galia After five -days storage at 20 C in fall 2007, line ASxAS fruit remained firmer when harvested at stage ZYG At stages ZG, HS and FS aroma and quality differences were few. On average, ASG fruit remained on the vine four days longer than Galia, suggesting a wider harvest window. Even though th ere were some differences in volatiles at stage ZYG, in order to enhance shipping quality, it is recommended that line ASxAS be harvested at stage ZYG where SSC was acceptable and fruit firmness was greatest at harvest and after storage

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16 CHAPTER 1 INTROD UCTION The Galia ( Cucumis melo L. var. reticulatus Ser.) muskmelon was developed in Israel by breeder Zvi Karchi and released in 1973. As a F1 hybrid, Galia has specific female and male parental lines. The female parental line of Galia is a Ha Ogen type melon cultivar called Noy Y izreel (Karchi, personal comm., 2004 ), which is g reen fleshed with a smooth, sutured skin ( Karchi, 2000). The male parental line of Galia was originally from the Peninsula of Crimea, in the Ukraine, and is a cultivar c alled Krymka, in which fruits were round with a golden, netted skin and light green, firm flesh (Karchi, personal comm., 2004). The resulting hybrid cross the Galia muskmelon ha s round fruits with an orange -netted skin and a green, soft textured flesh with a unique musky aroma, h igh soluble solids content (13 to 15 Brix) and is a high yielding cultivar (Karchi, 2000). As a result of marketing campaigns with Agrexco and sales at the British food chain, Marks and Spencer, Galia muskmelons became po pular all over Western Europe, except France (Karchi, 2000). Although it is a high quality fruit Galia muskmelon has some limitations. Besides being highly susceptible to powdery mildew ( Podosphaera xanthii (formerly Sphaerotheca fuliginea Schlech ex Fr. Poll.) ) (Mitchell et al. 2006 and 2007 a ), another main disadvantage is its short shelf -life (Mitchell et al. 2007a and 2007c ; Nuez -Palenius et al., 2005). Galia muskmelon s may last two to three weeks if harvested at a pre -slip stage and s to red at l ow temperatures (Aharoni et al., 1993). Exporters have increased shelf life by harvesting fruit at an immature (pre ripe/slip) stage ; however this leads to lower quality fruit (Fallik et al., 2001; Canliffe and Shaw, 2002; Pratt. 1971). In order to achiev e the best flavor, Galia must be harvested at full -slip. To address Galias

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17 limitations, breeders have created a whole new market class called Galia -type (GT) muskme lons (Karchi, 2000). These GT culti vars have improve d disease resistance/tolerance as well as an increased shelf life (Karchi, 2000; Mitchell et al, 2006 and 2007a). Unfortunately, although these GT cultivars are firm, many often lack the flavor, aroma, and high sugar content of the original Galia hybrid (Mitchell et al. 2006 and 2007a ). In order to attain an increased shelf life and maintain the high quality and flavor of the original Galia muskmelon, research ers have transformed the Galia male parental line (cv. Krymka) with an antisense ACC -oxidase gene (CMACO 1) (Nuez -Pale nius et al., 2006a). ACC oxidase is the catalyst in the last step in the biosynthetic pathway of ethylene (Yang and Hoffman, 1984), a hormone that has a major role in fruit ripening and senescence (Abeles et al., 1992). This hormone initiates fruit soften ing, changes in carbohydrate metabolism, aroma volatile production, and abscission, but does not regulate fruit size and sugar content (Pech et al., 1999; reviewed by Nuez -Palenius et al., 2008). The work from Nuez -Palenius et al. (2006a) produced two independent antisense Krymka lines named TGM -AS 1 and TGM -AS 2. Fruits from TGM -AS 2 line produced less ethylene and were firmer than untransformed fruits at half and full -slip stages (Nuez -Palenius et al., 2006b). TGM -AS 1 fruits also exhibited lower ethylene production, but only during the half -slip stage (Nuez Palenius et al., 2006b). Conversely, the female parental line of Galia muskmelon, cultivar Noy Yizreel was unable to be transformed with an antisense ACC -oxidase (CMACO 1) gene (Nuez Palenius et al., 2005).

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18 Nonetheless, an additional result of Nuez Paleniuss work was the development of antisense Galia muskmelon hybrids (T0GMH -AS 1 and T0GMH -AS 2) produced from the T0 transgenic male parental lines (TGM -AS 1 and TGM -AS 2) (Mitche ll et al., 2007c ). During fall 2004, T0 TGMH -AS 1 and T0 TGMH -AS 2 lines remained on the vine longer than Galia (Mitchell et al., 2007c ). However, in a previous crop, a severe powdery mildew ( P. xanthii ) epidemic led to no differences in days to harvest (DTH) between AS lines and Galia (Mitchell et al., 2007b). Due to these challenges, it was desirable to obtain AS Galia hybrids where both the male and female line incorporated the antisense ACC -oxidase (CMACO 1) gene. With both parents positive for t he transgene, it was hypothesized that the F1 hybrid progeny fruit would have reduced ethylene production and therefore a longer shelf -life. T he objectives of this research were three -fold. The first research objective was aimed at producing an elite st ock of the antisense male TGM -AS 1 and TGM -AS 2 lines provided by Nuez -Palenius and use these lines to produce an antisense ACC -oxidase (CMACO 1) female par ental line through backcrossing. A second objective was to produce antisen se Galia (ASG) hybrid muskmelon s through traditional breeding methods w here both the female and male parental lines possess the antisense ACC oxidase (CMACO 1) gene. The third and final objective was to ev aluate the ASG muskmelons by collect ing fruit quality data (fruit size, firmness, soluble solids content (SSC), titratable acidity, pH, and aroma) and ethylene and respiration rates These data were to be collected at different stages of growth to establish fruit quality characteristics of ASG muskmelons and to develop optim al harvest time guidelines. It is also important to note that because the fruit quality factor, aroma, had not previously been evaluated on

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19 muskmelon at the University of Florida Horticultural Sciences D epartment, aroma volatile collection and analysis procedures were determined. Aroma is an important fruit quality factor that is involved in fruit flavor and may be negatively affected by low ethylen e production since increased aroma coincides with fruit ripening. In order to accomplish these objectives research was conducted from 2004 through 2007 and is summarized in th e following chapters. Chapter 2 Literature Review, summa rizes the past and current literature that surrounds this research. Chapter 3 Production, Evaluation and Selection of Antisense ACC -oxidase (CMACO 1) Galia Muskmelon (Cucumis melo L. var. reticulatus Ser.) describes the d evelopment of elite parental lines, bearing an antisense ACC -oxidase gene. The elite parental lines were crossed in multiple combinations (AS AS antisense Galia ( ASG ) hybrid seed was produced. ASG hybrids then were evaluated and selections were made for additional studies. Chapter 4 Galia Muskmelon Fruit Quality and Flavor (Cucumis melo L. var. reticulatus Ser. ), describ es fruit quality factors and characterizes Galia and GT muskmelon aroma volatiles and flavor. Chapter 5 Aroma Volatile and Fruit Quality Evaluation of Antisense ACC Oxidase (CMACO 1) Galia Muskmelons (Cucumis melo L. var. reticulatus Ser. ), compares th e ASG and Galia muskmelons over three seasons, evaluating fruit quality, including aroma, bot h at harvest and after storage. And lastly, Chapter 6 Conclusions, summarizes the results and discusses the significant outcome of this work.

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20 CHAPTER 2 LITERA TURE REVIEW Introduction Classification and O rigin Cucumis melo L., commonly called melon belongs to the Cucurbitaceae or gourd family, a large family that consists of about 120 genera and over 800 species, including pumpkin and squash ( Cucurbita spp .) cucumber ( Cucumis sativa ) and watermelon (Citru llus lanatus ) (Whitaker and Davis, 1962; Jeffrey, 1990; Seymour and McGlasson, 1993; Nayar and More, 1998; Robinson and Decker -Walters, 1999) Within the Cucumis melo species there are numerous genetically di verse, polymorphic, interfertile subspecies or groups (Whitaker and Davis, 1962; Seymour and McGlasson, 1993; Nayar and More, 1998) These subspecies or groups have been analyzed and categorized by several scientists, including Naudin,1859; Whitaker and D avis, 1962; Smith and Welch, 1964; Munger and Robinson, 1991; Kirkbride, 1993; Robinson and Decker -Walters, 1999; Guis et al., 1998; and Pitrat et al., 2000. In 1859, Naudin divided Cucumis melo into ten groups. Eventually, they were regrouped into seve n (Whitaker and Davis, 1962; Smith and Welch, 1964; Guis et al ., 1998). These seven groups include variations of cantalupensis reticulatus inodorus saccharinus chito dudaim conomon and flexuosus The cantalupensis group, which is believed to have originated from Cantalupp e Italy (Robinson and Decker -Walters, 1997) are rarely grown in the U.S. and are true cantaloupes ( Whitaker and Davis, 1962) They are usually non-netted, ha ve a rough or warty skin with prominent sutures (Whitaker and Davis, 1962), and flesh is orange or

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21 green. However, the name (cantaloupe) is incorrectly attributed to netted melons in the U.S. (Bailey and Bailey, 1976). Reticulatus or muskmelon and also called rockmelons in Australia (Aubert and Bourger, 2004) have netted skin though some may have shallow sutures; flesh color ranges from green to deep orange ( Whitaker and Davis, 1962) These melons are sweet with a musky aroma (Whitaker and Davis, 1962). Inodorus or winter melon is a smooth -skinned fruit with minimal aroma, but has sweet white or green flesh and have a long shelf -life (Whitaker and Davis, 1962; Seymour and McGlasson, 1993). These include Canary, Casaba, Crenshaw and Honeyd ew (Seymour and McGlasson, 1993). Saccharinus melons are very sweet with smo oth skin that has green spots and some grey (Guis et al., 1998). Chito, or Mango melon or Garden Lemon are small, smooth and mottled fruits with an acidic flavor that are used as ornamentals or for pickling ( Whitaker and Davis, 1962). Dudaim or Pomegranate me lons are also sometimes grouped with chito (Naudin, 1859). These are small fruits with a yellow rind, white to pink flesh and have a musky odor (Whitaker and Davis, 1962; Guis et al., 1998). The most common type of dudaim is Queen Annes Pocket Melon, which has become localized in Louisiana and Texas (Whitaker and Davis, 1962). Conomon, also called Chinese cucumber or Oriental Pickling Melon, are small, oblong fruits with white, crisp flesh, a smooth exterior and have minimal aroma (Naudin, 1859; Guis et al., 1998; Whitaker and Davis, 1962).

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22 And Flexuosus (syn Utilissimus) is a long, serpent like melon used in salads in the IndoGangetic plains (Nayar and More, 1998). It is also called Snake Melon or Serpent Melon (Whitaker and Davis, 1962). T he cantal upensis reticulatus and saccharinus groups have been suggested to be grouped together (Munger and Robinson, 1991). Dudaim and chito have also been grouped as one under the chito name (Naudin, 1859; Munger and Robinson, 1991; Guis et al., 1998). However in 2000, Pitrat and his colleagues suggested only two groups: sweet melons, which included cantalupensis reticulatus inodorous and makuawa groups; and non-sweet, which included chate (chito ), flexuosus and conomon groups (Bu rgur et al., 2003 ). Nonethele ss, it remains debatable if the se groups serve any useful purpose except for horticultural characteristics (Whitaker and Davis, 1962). All groups of Cucumis melo easily hybridize together (Whitaker and Davis, 1968); and Cucumis melo is a cytologically s table diploid crop with 12 chromosomes (2n = 24) (Nayar and More, 1998; Doijode, 2001). Most muskmelons are herbaceous annuals, few are perennials, and all are frost sensitive ( Nayar and More, 1998; Bailey and Bailey, 1976). They are predominantly warm -se ason crops and are popular summer fruits (Nayar and More, 1998) and desserts (Bailey and Bailey, 1976). Origins stem from tropical and temperate subtropical areas of Africa, A sia and India, many of which thrive in hot and humid or desert conditions (Whitak er and Davis, 1962; Robinson and Decker -Walters, 1999; Munshi and Alvarez, 2005 ). Muskm elons can be tra ced as far back as 3000 B.C.E. and 2400 B.C.E. when the Ancient Egyptians cultivated and illustrated this fruit in paintings ( Robinson and Decker Walter s, 1999; reviewed by Walters, 1989; Mills, 2000). They are found in the wild

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23 throughout Africa and Asia, although truly wild forms and the largest concentration of non -cultivated species are found in Africa (Whitaker and Davis, 1962; Karchi, 2000). It is believed melons were introduced into Asia around 20001500 B.C.E. by way of the Silk Road (Karchi, 2000; reviewed by Walters, 1989), but culture of melons is thought to have started independently in both Africa and Asia, thus a center of origin is difficul t to identify (Kerje and Grum, 2000). As melon was domesticated in Africa and Asia it is thought to have then s pread into Europe and throughout much of the world ( Kerje and Grum, 2000). Reportedly, Columbus brought muskmelon seed to the New World on hi s second voyage, and it was grown on Isabella Island, of the Galpagos Islands (Robinson and Decker -Walters, 1999; Boswell, 2000). The Spaniards introduced it to California in 1683 (Robinson and Decker -Walters, 1999). There are records of production date d back to 1609 in Bermuda and 1650 in Brazil (Boswell, 2000). In 1796, the first cantaloupe seeds from Tripoli arrived in Philadelphia via Bernar d McMahon of Ireland (McDonald and Copeland, 1997). In 1881, a green-fleshed, netted ( reticulatus ) cultivar, Nette d Gem, was introduced by W. Atl ee Burpee Co. (Mills, 2000; Blinn, 1906). It is believed that many modern muskmelons are derived from Burpees introduction (Mills, 2000). One such derivative, Rocky Ford was originally from Rockyford, Co and was im portant in the commercial cantaloupe industry in the 1800s (Robinson and Decker -Walters, 1999; Blinn, 1906). Burpee Hybrid was the first melon F1 hybrid introduced in 1955 (Robinson and Decker Walters, 1999). Today, the U.S. classifies melons by area W estern and Eastern shipper -types, and specialty, which include Galia and other high quality melons suitable for niche markets (Shellie and Lester, 2004) The Western shipper

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24 types are grown in Arizona, California, and Texas for both domestic and export m arkets (Shellie and Lester, 2004). However, Eastern shipper types, which are grown throughout the Eastern U.S. are more perishable and are primarily used loacally (Shellie and Lester, 2004). Both the Western and Eastern types are of the reticulatus group. Specialty type melons consist of many diverse groups such as r eticulatus (Galia, Persian, and Ananas types) and cantal upensis (Charentais types) (Shellie and Lester, 2004). Another popular melon in the U.S. is the Honeydew ( inodorus) which has a longe r storage life than reticulatus and cantalupensis (Mills, 2000). Consumption and Production The U.S. ranks 4th in production of melons with 4.2 % of the world market, behind China ( >50%), Turkey ( 6.1% ) and Iran (4.4%) (Borriss et al., 2006). From 1990 to 2002, the United States saw a 27 % increase in melon consumption due to healthier eating year -round availability, economic growth and improved cultivars ( ERS/USDA 2003). Since 2002, melon consumption still remains high (Lucier and Dettman, 2008). Howe ver, melon production in the U.S. decr eased 17% from 1992 to 2004 (Borriss et al., 2006). In 2004, total U.S. cantaloupe production was valued at $300.6 million and honeydew melon at $89.7 million (Borriss et al., 2006). The U.S. is the largest importer of melons worldwide (Borriss et al., 2006). In 2004, the U.S. imported $117.3 million of melons from Mexico and Central America from December to April and exported melons valued at $91 million to Canada and Japan (Boriss et al., 2006). U.S. domestic melon production is from April to December in most states, with California, Texas and Georgia the top producers (Borriss et al., 2006; ERS/USDA, 2003). Florida is not among the top producers in melons, but it is the leading producer of watermelons in the U.S ( Boriss et al., 2006).

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25 Characteristics Cucumis melo L. is an indeterminate vine crop that has tendrils a nd a prostate or climbing nature that produces soft, herbaceous stems and branches ( Munshi and Alvarez, 2005 ). It is an andromono ecious species, first producing male (staminate) flowers on the main stem and later producing perfect (hermaphroditic) flowers on the lateral vines (Munshi and Alvarez, 2004; Whitaker and Davis, 1962). Although predominantly andromonoecious, there are some melon cultivars that are monoecious or gynomonoecious (Peterson et al., 1983; More et al., 1980; Munshi and Alvarez, 2005 ; Whitaker and Davis, 1962). Fruits are produced through cross -pollination by bees. One flower requires 10 to 15 bee visits for adequate pollination to occur (Mills, 2000). Anthesis occurs in the morning between 5:30 and 6:30 a.m. at temperatures between 22 29 C ( Munshi and Alvarez, 200 5 ; Nanpuri and Brar, 1966). Flowers are open for one day and pollen viability decreases as the day progresses (More an d Seshadri, 1998). After pollination, the ovary wall expands and develops into pericarp (fruit wall) with an exocarp (skin), mesocarp (flesh) and endocarp ( Munshi and Alvarez, 2005). Melon fruit s are classified as an indehiscent pepo with three locule sec tions or ovaries (Robins on and Decker Walters, 1999). Muskmelon fruit growth and development follows a sigmoidal growth curve (reviewed by Pratt, 1971). Fruit set of melon is cyclic; one to four fruit are set in each cycle (Mills, 2000). After initial fru it set, flowers abort for five to eight nodes and then set fruit again. Fruit are ready to harvest about 80 to 120 days after planting seeds. Seed s are borne internally in the locular cavity mucilage along the receptacle tissue (Mills, 2000) Seed are v iable at full maturity (Whitaker and Davis, 1962; Robinson and

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26 Decker -Walters, 1997). Tens to hundreds or more of seeds are produced per fruit and will remain viable for many years if stored in dry and cool conditions (Robinson and Decker -Walters, 1997; M ills, 2000; Doijode, 2001). Climacteric and N on -C limacteric Fruit Fruits had been classified either climacteric or non-climacteric based on their pattern of respiration levels during fruit ripening (Kidd and West, 1925). After the introduction of gas c hromatography, the plant hormone ethylene could be detected and was found to have a large increase during the climacteric in fruits (Burg and Thimann, 1959, 1960). Fruit ripening patterns that are climacteric exhibit a rise in respiration concurrent with an autocatalytic production of ethylene (Abeles et al., 1992) known as the climacteric peak (Seymour and McGlasson, 1993). Examples of climacteric fruits are apple (Malus domestica), avocado ( Persea americana ), banana (Musa ), and tomato (Solanum lycopersi cum ) (Kader, 2002a ). Non -climacteric fruits do not have a rise in respiration or ethylene during fruit ripening. Such non -climacteric fruits include citrus (Citrus ), cucumber (Cucumis sativus ) and strawberry ( Fragaria x anannasa) (Kader, 2002 a ). The ca ntalupensis and reticulatus melon groups are climacteric (Lyons et al., 1962; Seymour and McGlasson, 1993; Flores et al., 2002); and have a moderate respiration rate (10 to 20 mg CO2 kg1 hr1) and high ethylene production rate (10.0 to 2H4/ kg1 hr1) (Kader, 2002a ). M uskmelons and cantaloupes were reported to have a rapid climacteric at or near full maturity and abscission, with an interval from the pre climacteric to the climacteric peak being 24 to 48 hours (Lyons et al., 1962). However, mel ons of the inodorus group are nonclimacteric (Pratt et al., 1977) and have a low respiration rate (5 to 10 mg CO2 kg1 hr1) and produce moderate amounts of

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27 2H4 kg1 hr1) (Kader, 2002a ). Kendall and Ng (1988) reported high amounts of ethylene in netted melons ( reticulatus ) at or near harvest and while non netted melons ( inodorous ) did not produce et hylene until about 20 days postharvest. The rise in ethylene production during ripening is thought to control changes in color, aroma, texture and flavor (Lelivre et al., 1997; Hadfield et al., 1995; Alexander and Grierson, 2002; reviewed by Nuez -Paleni us et al., 2008). These changes have promoted extensive research on ethylene and fruit ripening (reviewed by Theologis, 1992; Shellie and Saltveit, 1993; Yamamoto et al., 1995; Lelivre et al., 1997; Hadfield et al., 1995; Alexander and Grierson, 2002; re viewed by Nuez -Palenius et al., 2008 ) and fruit quality (reviewed by Saltveit, 1999; reviewed by Nuez Palenius et al., 2008 ). There are both ethylene dependent and independent events associated with ethylene production (Pech et al., 1999; reviewed by N uez -Palenius et al., 2008). In Charentais cantaloupes ( cantalupensis ), ethylene production during melon fruit ripening were reported to stimulate carbohydrate metabolism, yellowing of the rind, fruit softening, respiration, aroma volatile production and abscission (Pech et al., 1999; reviewed by Nuez -Palenius et al., 2008). Ethylene independent activities include flesh color development, sugar and organic acid accumulation, loss of acidity and accumulation of 1 aminocyclopropane 1 -carboxylic acid (ACC) (Pech et al., 1999). In Krymka muskmelons ( reticulatus ), which are also the male parental line of Galia muskmelon, ethylene during ripening is associated with yellowing of the rind and fruit firmness loss while ethylene independent events are fruit wei ght and size, titratable acidity, seed number, mesocarp size ,+ and total soluble solids ( reviewed by Nuez Palenius et al., 2008). Ethylene production has also been associated with postharvest

PAGE 28

28 decay (Li et al., 2006) and chilling injury (Pech et al., 1999 ), thus reducing ethylene biosynthesis could be a method of reducing postharvest losses and extending shelf life in fruits ( reviewed by Nuez -Palenius et al., 2008). Ethylene biosynthesis follows the pathway from methionine via S adenosylmethionine (SAM) and 1 aminocyclopropane 1 -carboxylic acid (ACC). The enzyme responsible for catalyzing the conversion of SAM to ACC, is ACC synthase (ACS) and the enzyme catalyzing ACC to ethylene is ACC oxidase (ACO) (Yang and Hoffman, 1984). There are five ACS genes ( CMe -ACS1 to CMe -ACS5 ) that have been isolated from melon (Yamamoto et al., 1995; Ishiki et al., 2000; reviewed by Li et al., 2006; reviewed by Nuez -Palenius et al., 2008; reviewed by Ezura et al., 2008). CMe -ACS1 is expressed in the mesocarp tissue of ri pening fruit and is a wound responsive gene (Yamamoto et al., 1995). CMe ACS 2 is expressed in the fruit at the early stages of ripening (pre -climacteric) and in seedlings (Yamamoto et al., 1995). CMe -ACS 3 is also expressed in the mesocarp at the pre-climacteric stage, but at lower levels than CMe ACS 2 (Ishiki et al., 2000). The role of CMe -ACS 4 is not clear, but CMe -AC S5 is expressed in ripened fruit and is ethylene independent (reviewed by Li et al; reviewed by Nuez -Palenius et al., 2008 ; reviewed by Ezur a et al., 2008). There are three ACO genes in melon (reviewed by Li et al, 2006; reviewed by Nuez -Palenius et al., 2008 ; reviewed by Ezura et al., 2008 ). CM ACO1 is expressed in etiolated hypocotyls, roots, leaves flowers and fruit; and it has a role i n fruit ripening and senescence as well as in response to wounding (Lasserre et al., 1996; Guis et al., 1997; reviewed by Li et al., 2006; reviewed by Nuez Palenius et al., 2008 ; reviewed by Ezura

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29 et al., 2008 ). CM -ACO 2 is expressed in etiolated hypocotyls and CM -ACO 3 is expressed in many tissues, but not in fruit (Lasserre et al., 1996; Guis et al., 1997; reviewed by Li et al ; reviewed by Nuez Palenius et al., 2008 ; reviewed by Ezura et al., 2008). The suppression of ethylene via blocking ACS and/or ACO in fruits has resulted in inhibition of the ripening process (reviewed by Pech et al., 2008). In higher plants, there are two systems of ethylene production. System I occurs in all vegetative tissues and in the fruit before it ripens; and produces basa l ethylene levels (reviewed by Jakubowicz, 2002; Alexander and Grierson, 2002). System II occ urs once the fruit reaches its climacteric ph ase, when ethylene production is autocatalytic and during petal senescence (Alexander and Grierson, 2002; reviewed by Jakubowicz, 2002). Not only is ethylene considered the ripening hormone, but it is also responsible for many other physiological and developmental processes such as adventitious root formation, flower opening, leaf/flower senescence/abscissio n, seed ger mination and responses to both biotic and abiotic stresses (Yang and Hoffman, 1984; Abeles et al., 1992; Yang and Oetiker, 1998; Saltveit, 1998 ; reviewed by Ezura et al., 2008). Essentially, the effect of ethylene on the fruit ripening process prepares th e fruit for seed dispersal, by facilitating the visual and olfactory responses that make the fruit attractive to animals that will assist in releasing the seed (reviewed by Nath et al., 2006; Adams -Philips et al., 2004 ; reviewed by Ezura et al., 2008). Mat urity and Ripening Fruit quality and shelf -life is dependent on maturity at harvest. Fruits that are harvested either immature or overripe have poor quality and subject to damage (Kader 2002b ). The maturation of melons depends on the fruit type ( Seymour and McGlasson, 1993). As muskmelons mature and ripen they develop a full net, the background rind

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30 color will change and an abscission zone will develop (Shellie and Lester, 1996). The fruit will begin to separate from the stem at the abscission zone as i t becomes fully ripe Fruits are fully ripe when the stem separates completely from the fruit, also called full slip stage (Sykes, 1990; George, 1999). However, most muskmelons are commonly harvested when the stem separates half way, or half -slip sta ge (Kasmire et al., 1970). Muskmelons harvested at peak of ripening will have high soluble solids and produce aroma while those picked too early ( prior to abscission zone development ) will have reduced soluble solids, aroma and flavor, and those harveste d too late (overmature) will have reduced quality (Seymour and McGlasson, 1993; reviewed by Pratt, 1971; Rosa, 1928; Lloyd, 1928). Generally, muskmelons are ready to harvest at about 42 days after anthesis (Agblor and Waterer, 2001). Inodorus types, such as honeydew, do not slip from the vine until they are overripe and instead, are harvested by cutting the melon from the vine (Pratt et al., 1977; Seymour and McGlasson, 1993). There are many types of melons with characteristics between that of muskmelon and honeydew, often making it difficult to determine the optimum harvest time ( Seymour and McGlasson, 1993). Fruit Q uality Since melons are often consumed as a dessert, optimum fruit quality is essential (Lloyd, 1928). Quality can be divided into both in ternal (aroma, flavor, texture) and external (color, firmness size ) factors, which changes depending on ones perspective (Shewfelt, 1999; Cause et al., 2003; Wills et al., 2007). In general, optimum fruit quality depends on fruit flavor which comprises the organoleptic attributes of taste (including fruit sweetness and acidity), aroma, texture, and color (Yamaguchi et al., 1977; Li et al., 2006; Goff and Klee, 2006; Causse et al., 2002). Flavor is perceived predominantly by

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31 aroma receptors in the nose a nd taste receptors in the mouth (Fisher and Scott, 1997). Dependent on many variables, flavor is th erefore a complex trait determined by many factors including culture, environment, genetics, production, and postharvest handling (Baldwin, 2002). Sweetne ss Fruit sweetness has been determined to be the most important fruit quality component (Yamaguchi et al., 1977). In the U.S. melons with a soluble solids content (SSC) of 9 Brix are considered grade No. 1 and those with a SSC of 11 Brix or higher a re grade Fancy (Lester and Shellie, 2004). Sucrose is the dominant sugar in ripe fruits, while glucose and fructose are the dominant sugars in fruits during the first 24 days after anthesis (McCollum et al., 1988). Since muskmelons do not have a starch reserve and therefore, no carbohydrates to convert to sugar, they will not increase in so luble sugars after harvest; thus, it is essential to harvest muskmelons at their optimum maturity (reviewed by Pratt, 1971; Burger et al., 2006). T he longer the frui t remains on the plant accumulating sugars, the higher the SSC (Bianco and Pratt, 1977). Sucrose accumulation in melons is regulated by the inverse relationship of the enzymes, sucrose phosphate synthase (SPS), which accumulates during melon development a nd acid invertase (AI), which decreases (Burger and Schaffer, 2007; Lester et al., 2001; Hubbard et al., 1991; Hubbard et al., 1990; Hubbard et al., 1989; McCollum et al., 1988; Schaffer et al., 1987; reviewed by Seymour and MacGlasson, 1993; reviewed by N uez -Palenius et al., 2008). In a study that evaluated both sweet and non -sweet melon cultivars, the enzymes, SPS, sucrose synthase and neutral invertase were also positively correlated with sucrose accumulation in all melon types, though there was low ac tivity of these enzymes in the non -sweet melons types (Burger and Schaffer, 2007). Although sucrose accumulation is

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32 associated with the biochemical processes involved in ripening, it is also related to cultural factors (reviewed by Pratt, 1971). Fruits ca n obtain a high SSC with low night temperatures, a longer maturation period, large leaf area, and harvesting fully ripe fruits (Welles and Buitelaar, 1988). Conversely, low SSC can be a ttributed to harvesting fruit too early, such as prior to development of the abscission layer ( Seymour and Mcglasson, 1993). Acids Acids, though of high importance in many other fruits such as tomato (Solanum lycopersicum ) (Burger et al., 20 03; Cause et al., 2003), have minimal effect on melon fruit quality (Yamaguchi et al ., 1977). Sweet melons ( reticulatus cantalupensis inodorus and makuwa groups) have a low organic acid content, with citric acid being the major organic acid (reviewed by Burger et al., 2006; Burger et al., 2003; Leach et al., 1989). Malic acid is also present in the mesocarp, but at much lower levels than citric acid (reviewed by Burger et al., 2006). Titrable acidity (% citric acid) in various melon types can range from 0.054% in Galia muskmelons to 0.138% in Tendril winter melons, with no differences in pH (range: 5.74 to 5.65) (Artes et al., 1993). Texture Melon firmness or texture is a critical characteristic since it directly relates to the postharvest shelf life, transportability and pathogen susceptibility of the fruit (reviewed by Li et al. 2006). Texture preferences vary among consumers, as m uskmelons that received poor ratings in sensory panels have been described as being too soft as well as too hard (Yamaguchi et al., 1977 ; Pardo et al., 2000). Nonetheless, i t is considered to be the third most important attribute (fruit sweetness is first, followed by aroma and/ or flesh color) that affects eating quality in muskmelons (Yamaguchi et al., 1977). Pectins are the

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33 primary component in the cell wall contributing to fruit texture (reviewed by Prasanna et al., 2007). Muskmelon fruit softening is not clearly defined as it could be the result of ethylene production during ripening (Lester and Dunlap, 1985; Pratt et al., 1977; Bianco and Pratt, 1977) and/or the result of cell wall degradation d ue to enzymes (Rose et al., 1998; Seymour and McGlasson, 1993; reviewed by Nuez -Palenius et al., 2008). W ater loss after harvest is another factor that may contribute to fruit softening (Lester and Bruton, 1986) a s well as the loss of mesocarp membrane i ntegrity as determined by high electrolyte leakage (Lester, 1988). Melon softening during fruit ripening has also been related to changes in pectic and hemicellulosic polysaccharides as well as a net loss of non -cellulosic neutral sugars (McCollum et al., 1989). In Charentais cantaloupe fruits, there are also modifications in pectic and hemicellulosic polymers during ripening; and changes in the hemicelluose, xyloglucan were closely associated with cellulose microfibrils that may have an affect in early softening (Rose et al., 1998). The enzyme polygalacturonase (PG) is also attributed to pectin disassembly in melon (Hadfield et al., 1998), though the role and presence of PG in melon has also been disputed (McCollum et al., 1988, 1989; Lester and Dunlap 1985; reviewed by Pratt, 1971). Color Flesh color in melon varies among types, contributing to their uniqueness and importance in fruit quality (reviewed by Pratt, 1971; Yamaguchi et al., 1977). Flesh pigments in melon can be many colors and vary from orange, light orange, pink, green, and white (reviewed by Nuez -Palenius et al., 2008) or magenta/red -fleshed (Mitchell Harty et al., 200 9 a). In o range fleshed melons the predominant pigment is beta carotene though other pigments found include alpha -car otene delta -carotene, lutein, phytofluene phytoene, violaxantin, xanthophylls and traces of other carotenoids (reviewed by Pratt,

PAGE 34

34 1971; Watanabe et al., 1991). Flesh coloring begins in the center of the fruit and progresses outward until color is unifor m at maturity (Reid et al., 1970). Carotenoids have been detected in muskmelons as young as 10 days, but visual detection did not occur until 20 days post anthesis (Lester and Dunlap, 1985). At 30 days post anthesis, a two to three -fold increase in beta carotene levels was also observed (Lester and Dunlap, 1985). And as carotenoids accumulate during melon development, chlorophyll content decreases (Reid et al., 1970; reviewed by Pratt, 1971). In a study of both light orange and orange flesh melons, the l ight orange flesh melons had about 50% less beta -carotene than the orange flesh types while green and white flesh melons had the leas t amount of carotenoids (Watanabi et al., 1991). Aroma The characteristic flavor that is associated with melons is also d ependent on its aroma. Aroma volatiles are released as the fruit ripens and their presence, absence and quantity characterize each melon type (Pratt, 1971; Teranishi, 1971 ; reviewed by Engel et al.,1990). According to Pratt (1971) research on muskmelon ar oma began in the 1930s when Rakitan (1935, 1945) identified and noted increases during ripening of the compounds acetaldehyde and ethanol in muskmelons. In 1957, the compounds acetoin and 2, 3 butylene glycol were measured in muskmelon and revealed diffe rent volatile patterns. Acetoin was detected in fruits when they were over -ripe, while 2, 3 butylene glycol was found at harvest and increased until ideal eating stage, but disappeared when they were over ripe (Serini, 1957 reviewed by Pratt, 1971). Today, over 240 aroma compounds have been identified in muskmelon and the predominant aromatic compounds are esters, alcohols and aldehydes (Obando -Ulloa et al., 2008; Beaulieu, 2006; Lamikanra, 2002; Beaulieu and Grimm, 2001 and 2002;

PAGE 35

35 Nijssen and Visscher, 199 6 ; Njissen et al., 1996). Esters have a major role in fruit flavo rs, and are responsible for most of the flavor in melons and many other fruits (Fisher and Scott, 1997). Alcohols are less important to flavor, as they have a higher odor threshold value (F isher and Scott, 1997), but play an important role in melon aroma (i.e. cis 6 -nonen1 -ol, a green, melon aroma compound (Kemp et al., 1972, 1973, 1974). Aldehydes are also responsible for fruity aromas and may provide a characteristic flavor (Fisher and S cott, 1997). The a roma volatiles that are considered to be the most important compounds are determined by their odor value (OV) (also called log odor units) which is the ratio of the concentration of the compound to its know n odor threshold value (OTV) (Bauchot et al., 1998; Berger, 1995; Buttery 1993; Teranishi et al., 1991). When the OV of a compound is greater than 1.0 that compound is considered to be a significant contributor to the overall aroma ( Bauchot et al., 1998; Berger, 1995; Buttery 1993; Teranishi et al., 1991) Odor threshold values (OTVs) are usually determined in air, water or mineral oil by several investigators with different test methods; thus OTVs can vary by as much as 1000 (Fischetti, 1994). Aroma volatiles are believed to b e under genetic control as there are noticeable differences between melon cultivars ( Yahyaoui et al., 2002; Wyllie and Leach, 1992; Wyllie et al., 1996a ). Seeds can also have an influence on aroma as pollinated fruits have a favorable aroma over parthenoc arpic fruits probably due to higher carbohydrate levels in pollinated fruits (Li et al., 2002). And higher amounts of aroma volatile compounds have been reported in mature cantaloupes compared to fruits at immature stages (Beaulieu, 2006; Senesi et al., 2005; Beaulieu and Grimm, 2001; Wang et al., 1996; Horvat and Senter, 1987; Yabumoto et al., 1977 ).

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36 In other early studies of muskmelon aroma, over 50 volatile compounds were identified in muskmelons (Kemp et al., 1971, 1972a 1973; Kemp et al., 1972b). The compounds, cis 6 -nonen1 ol and cis 6 nonenal were associated with the muskmelon like, musky or green melon aroma (Kemp et al., 1972, 1973, 1974). However, Yabumoto et al. (1977) suggested that dimethyl disulfide and other sulfur compounds may also be important comp onents in muskmelon fruit flavor. This was also confirmed by Wyllie and Leach (1992) and Wyllie et al. (1993) who evaluated additional sulfur compounds. Sulfur compounds have a high impact on flavor since they bind to olfactory receptors (Fisher and Scot t, 1997). Yabumoto et al., (1978) reported that large quantities of volatile esters were found to be critical for the characteristic fruity aroma in muskmelon and that there were at least three different volatile groups (acetaldehyde and ethanol; ethyl est ers; and acetate esters), that could be determined based on their specific patterns; and that acetate esters increased rapidly and plateaued while the other two groups exhibited a continuous accelerating rate of production. Differences in aroma were also o bserved in two cantaloupe cultivars, where PMR 45 was more aromatic than Top Mark (Yabumoto et al., 1978). Yabumoto et al. (1978) concluded that the more aromatic PMR 45 was attributed to its higher ethylene production than Top Mark. An additional eight compounds were identified by Horvat and Senter (1987) from cantaloupes at different maturity stages Schieberle et al. (1990) concluded that the major aromatic compounds of muskmelon were methyl 2 -methylbutanoate, ( Z ) 3 -hexenal, ( E ) 2 hexenal and ethyl 2 -methylpropanoate. Wang et al. (1996) reported that ethanol was the only volatile in immature muskmelons whereas other volatiles such as ethyl acetate

PAGE 37

37 and 2 -methylbutyl acetate developed after 32 days post -pollination. Esters are considered to be positive aroma compounds and alcohols negative due to their fermented note (LoScalzo et al., 2001). Senesi et al. (2002) also concluded that ethyl esters such as ethyl acetate were highly correlated with high total aroma and consumer acceptance; and t h erefore, could be a marker of an optimum -quality melon. Also, the method that volatiles are measured is another factor to consider when determining aromatic profiles (Jordan et al., 2001). Volatile collection from muskmelon essence and fruit puree demo nstrated differences in compound in detection (Jordan et al., 2001). Aroma volatiles vary not only among different melon cultivars (Senesi et al., 2002), but also among different melon types such as honeydew melons ( Cucumis melo L. var. inodorous ), whic h have at least three unique compounds, ( Z ) 6 -nonenyl acetate, ( Z ) 3 nonenyl acetate and ( Z, Z ) 3, 6 -nonadienyl acetate compared to other melons (Buttery et al., 1982). Aroma volatiles from cultivar Golden Crispy melons, which have a smooth yellow skin a nd white flesh, revealed the presence of thioether esters and dioldiesters, not previously reported in melons (Wyllie and Leach, 1990). Queen Annes Pocket melons ( Cucumis melo L. var. dudaim ) are noted for their aroma, not taste, which corresponded to h igher levels of volatiles found in the skin rather than the pulp (Aubert and Pitrat, 2006). True cantaloupes ( Cucumis melo L. var. cantalupensis ) are another melon type where over 100 volatile compounds, which include sulfur compounds such as 2 (methylth io)ethanol have been reported (Homatidou et al., 1992). Another type of cantaloupe, Charentais ( Cucumis melo L. var. cantalupensis ), are prized and for their

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38 aroma and sweet flavor (Aubert and Bourger, 2004; Goldman, 2002). M ore than 80 compounds have be en identified in Charentais melons, with the esters, ethyl acetate, 2 methylpropyl acetate and 2 -methylbutylacetate comprising 60% of the total identified volatiles (Bauchot et al., 1998). However, aroma development in Charentais cantaloupes that have bee n transformed with an antisense ACC oxidase gene had a significant decrease in volatiles, having only 20 30% of the total quantity of acetates as compared with nontransformed fruit (Bauchot et al., 1998 and 1999). Ethylene, therefore is also believed to have an effect on aroma, as the ethylene inhibited melons demonstrated reduced aroma (reviewed by Zhu et al., 2005; Bauchot et al, 1998 and 1999). This reduction in aroma volatiles could be due to the reduction in esters, which are catalyzed by an alcohol acetyltransferase (AAT) enzyme, which is regulated by ethylene (Bauchot et al., 1998). Analysis of different Charentais cantaloupe cultivars with wild, mid and long storage shelf life concluded that long shelf life (LSL) types were the least aromatic, p robably due to their lower ethylene production (Aubert and Bourger, 2004). Saftner et al. (2006) also reported higher volatiles in cantaloupe (climacteric) versus honeydews (non -climacteric). Aroma volatiles have also been studied on various Galia ty pe cultivars (Leach et al., 1989; Wyllie and Leach, 1992; Fallik, et al., 2001 and 2005 ; Hoberg et al., 2003; Obando-Ulloa et al., 2008; Shalit et al., 2001; Kourkoutas et al., 2006). Wyllie and Leach (1992) concluded that sulfur -containing compounds in t he aroma volatiles of muskmelon were important and found that the Galia muskmelon used in their study had relatively intense amounts of 2 (methylthio)ethyl acetate. Research on Galia type muskmelon aroma found that in the GT cultivars, C8 and 5080 butyl acetate, 2 -

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39 methylbutyl acetate and hexyl acetate were the most abundant compounds (Fallik et al., 2001). Fallik et al. (2001) also reported how higher aroma does not necessarily imply higher consumer preference. Cultivar C8 had stronger aroma t han 5080, but contained less sugar and was not preferred as often as 5080 by a taste panel. Shalit et al. (2001) reported that volatile acetates were higher in ripe Arava fruits than in unripe ones; and that volatile acetates were correlated with to tal soluble solids. Volatile esters are formed via the enzymes, alcohol acetyltransferasaes (AAT) (EC 2.3.1.84), which catalyze the reaction between acyl CoA and alcohol during ripening ( El -Sharkawy et al., 2005). Khanom and Ueda (2008) reported that the esters isobutyl acetate and benzyl acetate were produced the most in melons (cv. Earls faveorite and cv. Prince). Shalit et al. (2001) found that AATs were present in Arava fruits and in creased as they ripened. This was in contrast to a non -climacteric, casaba type melon, which was a volatile acetatelacking nonaromatic melon with negligible AAT activity. Hoberg et al. (2003) did not measure actual quantities of aroma, but their studies with sensory panels reported that fruits with negative charac ters such as greeny, solvent and unpleasant odors were not preferred by consumers and could be differentiated from pleasant or fruity odors on the basis of retro nasal odor. Hoberg et al. (2003) concluded that cultivars C8 and Ideal had more n egative characters, espe cially after 16 days in storage; however, they are no longer grown in Israel as a result of their findings. Obando-Ulloa et al. (2008) reported that cultivar Fado was highest in propyl acetate, methyl 2 -methylbutanoate and hexyl acetate. Kourkoutas et al. (2006) found Galia contained higher levels of the acetate esters isobutyl, butyl, 2 -methylbutyl and hexyl acetate, than cantaloupe and honeydew.

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40 Although there has been research on Galia type muskmelon aroma, there are no known reports on the aroma of t he original Galia muskmelon. A lthough work by Leach et al, 1989 and Wyllie and Leach (1992) may have been with the original Galia cultivar, however they did not specify so and the work by Kourkoutas et al. (2006) report ed results on Galia muskmelons with no report of the actual cultivar name. Taste and Sensory A nalysis Organoleptic quality involves not only color, texture and aroma, but also taste (Causse et al., 2002). There are five major taste sensations: saltine ss, sweetness, sourness, bitterness and umami (savory) (Fisher and Scott, 1997). Taste sensations occur upon contact with food in the mouth (Fisher and Scott, 1997). Th erefore sensory measurements of quality attributes can also provide an approximation o f consumer acceptability (Abbott, 1999). Since muskmelons tend to be quite variable in quality, sensory analyses are another method used to determine muskmelon fruit quality (Aulenbach and Worthington, 1974; Yamaguchi et al., 1977). Sensory panels can be performed with both trained and untrained panelists (Fisher and Scott, 1997). Trained sensory panels can be expensive and slow (Studman, 2001), but information on both consumer satisfaction as well as identification of organoleptic qualities of the produ ct can be obtained (Gurineau et al., 2000). There are different types of sensory panels that include discrimination tests where a triangle test, paired comparison test, duo -trio test, and ranking test may be used; or affective tests (hedonic tests), whi ch assess consumer preference and/or acceptance and rate samples according to a hedonic category scale (Fisher and Scott, 1997). Panelists can be asked to rate and comment on a variety of quality attributes including: firmness/texture, flavor, sweetness, appearance, acceptability, juiciness and eating quality (Senesi et al, 2002; Saftner et al., 2006).

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41 Galia Muskmelon Galia and Galia Type Specialty C ultivars Developed in Israel in 1973 by plant breeder Zvi Karchi, the Galia muskmelon (Cucumis mel o L. var. R eticulatus Ser.) was the first F1 hybrid melon developed in Israel. It was the product of Noy Yizreel and Krymka cultivars. Noy Yizreel was a HaOgen type, which was a common Hungarian open-pollinated cultivar grown throughout Israel a nd valued for its high yield and quality. Krymka was an ear ly producing Ukrainian cultivar, which was selfed for ten generations and selected for uniformity ( Karchi, 2000; Karchi, personal communication, 2004). The result of this cross, the Galia mus kmelon, was a superior quality fruit with sweet, green flesh, a yellow netted exterior and a fragrant, musky aroma (Karchi, 2000). After its introduction, Galia muskmelon quickly became a popular new market name throughout Europe and the Mediterranean by way of an intense marketing campaign with Agrexco and sales at the popular British food chain, Marks and Spencer (Karchi, 2000) By the 1980s, Galia muskmelons were sold all over Western Europe, except France (Karchi, 2000). Its popularity is attributed to its intense flavor, aroma and sweetness. In effect, Galia F1 hybrid melon production was one of the factors that helped revive Israels agricultural production, breeding and research as well as increase its competive advantage in world markets (Karchi, 2000). Although it is a high quality melon, Galia muskmelon has some limitations. Besides being highly susceptible to powdery mildew ( Podosphaera xanthii ) (Mitchell et al. 2006 and 2007), another main disadvantage is its short shelf -life (Mi tchell et al. 2007a and 2007b; Nuez -Palenius et al., 2005). Galia muskmelons may last two to three weeks if harvested at a pre -slip stage and stored at low temperatures (Aharoni et al.,

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42 1993). In order to achieve the best flavor however, Galia must be harvested at full -slip. Exporters have increased shelf life, but decreased quality by harvesting fruit at an immature stage; however this leads to lower quality fruit (Fallik et al., 2001; Cantliffe and Shaw, 2002; Pratt. 1971). As a result of Galias limitations and its popularity, breeders have worked to improve disease resistance/tolerance as well as improve shelf life ( Mitchell at al., 2007). Today Galia is a trade name for o ther look a like melon cultivars, commonly called Galia type melons. Unfortunately, although these Galia type cultivars are firm, they often lack the flavor, aroma, and high sugar content of the original Galia hybrid (Mitchell et al., 2006, 2007a and 2007b) According to a survey of seed companies throughout the world, today there are over 75 Galia type muskmelons. At least 60 Galia types have been released as cultivars (Table 1). E ight seed companies currently sell the original Galia F1 hybrid (Bakker Brothers, Genesis, Golden Valley Seed, Hazera, Seeds of Chang e, Thompson and Morgan and Zeraim Gedera ). Of these eight companies, it is known that Hazera, Genesis and Zeraim Gedera actually produce the Galia F1 hybrid. A t least two of the companies, Golden Valley Seed and Thompson and Morgan purchase the seed fro m an outside source and re -sell it (J. Harty, survey, 2008). Galia M uskmelon P roduction Galia muskmelon is grown primarily in Israel, Morocco, Turkey, and Spain, being exported principally to the U.K. and Europe where it is in high demand (Rodriguez et al., 2002). Galia muskmelons grown in the Mediterranean for export to Europe usually weigh 0.9 to 1.5 kg and can have a SSC up to 14 Brix (Rodriguez, 2003). Galia muskmelons are also produced in Central America (Guatemala/Honduras) and some parts of U.S. (J. Ortiz, personal comm., 2008). The Galia muskmelon can be found in U.S.

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43 markets, but quality is low due to production and harvesting practices (Rodriguez et al., 2002) Generally, Galia muskmelons are field -grown and imported from countries w here it is picked immature, at a zero -slip stage, to ensure a longer shelf -life (Cantliffe and Shaw, 2002). The Galia cultivar is especially adapted to intensive irrigation and fertilization where yields of up to 50 Mt/ha of high quality fruits (13 to 15 Brix) have been recorded under arid environments (Karchi, 2000). During the 1970s, Galia muskmelon was produced in northern Israel under dry land farming conditions where nitrogen (N) was only applied prior to rainfall during the winter (Rodriguez et al, 2005). Production in southern Israel used different types of irrigation, which included rain -fed cultivation, complimentary and complete irrigation; and also required N applications throughout the season, depending on the crop growth stage (Hecht, 1998). Producers in the north of Israel grew under open field conditions while producers in the south grew under protected tunnels (D. J. Cantliffe, personal comm., 2008). Although Galia muskmelon is adapted for intensive field cultivation, production i n the Arava desert in Israel under protected structures, such as tunnels, is also used to protect crops from wind, rain and low temperatures (Rodriguez, 2003). Galia muskmelon production under protected structures is also employed in the U.S. with the u se of passively -ventilated tunnels and greenhouses (Jett, 2004; Shaw et al., 2001; Rodriguez et al., 2002; Rodriguez, 2003; Waldo et al., 1997 and 1998). Galia muskmelons are sensitive t o rainfall during flowering and fruit set, making them difficult to produce in the field in places such as Florida (Rodriguez et al, 2002). Rainfall is a lso a problem at harvest time, as i t has been found that SSC can decrease in some muskmelons

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44 after rain (Wells and Nugent, 1980; Bouwkamp et al., 1978). T h erefore, the pr otected structures are a method to ensure highest quality Galia muskmelons (Rodriguez et al, 2002). Production of Galia muskmelons in passivelyventilated greenhouses is accomplished by trellising plants in an upright vertical fashion and careful pruning of lateral shoots (Shaw et al., 2001; Rodriguez, 2003). Plants are grown in pots or bags with a soilless media and, twine, strung down from a steel cable, is clipped under the cotyledons and the plant is twisted up and around the twine as the plant g rows (Rodriguez, 2003). The soilless media used can be a variety of types, including composted pine bark and perlite (Rodriguez et al., 2006). During the first three to four weeks after planting, or the vegetative stage, all lateral branches are pruned up to the eighth node to allow for optimum plant growth that will improve fruit load support (Shaw et al., 2001; Rodriguez, 2003). After this stage, female flowers develop on the lateral branches and are pollinated via bumble bees ( bombus impatiens ) (Shaw e t al., 2001; Rodriguez, 2003). Pruning off lateral branches that do not set fruit as well as trimming lateral branches that have developing fruits (leaving only one or two leaves to serve as a source of assimilates) continues throughout the season (Rodriguez, 2003). Plant spacing in protected culture is important as growers make efficient use of production space in order to help off -set high greenhouse costs (Rodriguez et al., 2006). Studies of Galia type muskmelons planted at densities of 1.7, 2.5, 3.3 and 4.1 plants m2 demonstrated that yield increased linearly with increasing plant density, without negatively affecting fruit quality (Rodriguez et al., 2006). Therefore if market prices for Galia muskmelons are $1.44 per kg, plants grown at a density of 3.3 plants m2 will

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45 have almost double the returns than those at a density of 1.7 plants m2 (Rodriguez et al., 2006). Irrigation and fertilization of soilless grown muskmelons is accomplished through drip irrigation (Shaw et al., 2001). Just as Galia muskmelon production in southern Israeli soils required applications N throughout the season, it is also important in soilless production (Rodriguez et al., 2005). Adjusting N concentrations in the fertilizer solution according to different growth stages (flowering, flowering to fruit set, fruit development, and fruit ripening) is recommended to reduce over -fertilization (Rodriguez et al., 2005). Excess N causes plants to become more vegetative and reduces the potential to maximize fruit set (D. J. Cantliffe, personal comm., 2008). Galia Muskmelon Postharvest Practices In order to achieve peak flavor, Galia must be picked at a fully ripe or full -slip stage (Karchi, 1979; Mitchell et al. 2007a; Nuez -Palenius et al., 2005), which reduces storag e life. Exporters have increased shelf life, but decreased fruit quality by harvesting at an immature stage (Fallik et al., 2001; Cantliffe and Shaw, 2002; Pratt. 1971). Although there are Galia type (GT) cultivars available that have an extended shelf -life, they often lack the flavor, aroma and high s oluble solids content of the original Galia hybrid (Mitchell et al., 2007a). In addition to production of GT cultivars, several methods have been used to extend the postharvest shelf life of Galia and GT muskmelons. They can be harvested early, at a green, pre-slip or half -slip stage when fruits are firmer, but this often results in reduced sweetness and flavor (Fallik et al., 2001; Cantliffe and Shaw, 2002; Pratt. 1971). Muskmelons can also be stored at low temperatures (2.5 to 5 C) to maintain firmness (Asghary et al., 2005) or rinsed with hot water to reduce both fruit softening and decay

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46 development (Fallik et al., 2000; Lalaguna, 1998; Teitel et al, 1989). Galia muskmelons subjected to < 1.0 kGy of irradiation combined with a hot -water dip protected fruit from decay and did not affect quality (Lalaguna, 1998). Waxes have also been used to maintain internal and external melon fruit quality (Fallik et al., 2005; Aharoni et al., 1992). S odium bic arbonate has been reported to reduce decay as well as maintain firmness (Aharoni et al., 1997). Moreover, a combination of hot water and a wax treatment with sodium bicarbonate may also be used to reduce decay and increase 2007). Other postharvest treatments of GT muskmelons have included applications of hydrogen peroxide or treatments of thujaplicin a chelating agent that inhibits microbial enzymes ) (Aharoni et al., 1994; Aharoni et al., 1993). Storage of Galia muskmelons in a controlled atmosphere of 10% CO2 and 10% O2 both with and without an ethylene absorbent decreased fruit softening and deca y (Aharoni et al., 1993). T he use of 1 -methylcyclopropene (1 -MCP), an ethylene action inhibitor, suppressed softening of Galia muskmelons at both green and yellow maturity stages (Erg un et al., 2006). Although these methods can be effective in extending the postharvest shelf -life of melon, many result in an extra step that producers and distributor s would most l ikely choose to avoid. Th erefore instead of further complicating the postharvest handling process another means could be to modify the innate quality of the fruit itself To accomplish this, it is necessary to understand key features of the fruit The Galia muskmelon is a climacteric fruit I t has a burst of respiration concurrent with an autocatalytic production of ethylene (Abeles et al., 1992; Seymour and McGlasson, 1993). This is a defining feature of ripening in fruits such as melons

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47 (Bower et al ., 2002) and causes the fruit to ripen, abscise and soften very quickly (Abeles et al.,1992). This is why Galia has a short shelf life. Galia are best when harvested fully ripe, when the climacteric peak and abscission occur (Cantliffe and Shaw, 2002; Mitchell et al., 2007c) T herefore, knowledge about the ethylene biosynthetic pathway is important since it is essential to the fr uit ripening process. Furthermore, the last step in the ethylene biosynthetic pathway can be inhibited using antisense tec hnology, which blocks ethylene production in the fruit, therefore, making the fruit firmer (reviewed by Nuez Palenius et al., 2008) This has been accomplished in tomato (L ycopersicon e s culentum ) (Hamilton et al., 1990), Charentais and Vedrantais can taloupes (Ayub et al., 1996; Guis et al, 1997; Guis et al, 2000; Silva et al., 2004) and plums ( Prunus domestica L.) ( Callahan and Scorza, 2007). In addition, the most recent work using antisense technology, by Nuez -Palenius et al. (2006a), was able to transform the male parental line of Galia muskmelon (cv. Krymka) with an antisense ACC oxidase gene (CMACO 1). Antisense Krymka fruits produced less ethylene and were firmer than wild type (WT) fruits, yet soluble solids content (SSC) was similar to WT fruits (Nuez Palenius et al., 2006b). The work from Nuez -Palenius et al. (2006a and 2006b) provided an essential step towards the goal of improving the shelf -life, while maintaining the high quality and flavor of the original Galia muskmelon.

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48 Ta ble 2 1 List of Galia type musk melon (Cucumis melo L. va r reticulatus Ser .) cultivars. Seed Company Cultivar Name BakkerBrothers Antalya, Bardot, Bastogne D. Palmer Seed Gala Girlie, GM 04 22, Sigal De Ruiter Abellan, Ajax, Cyro, Riscal, Supra, Zo ndra Emerald Seeds Melon EM 740 F 1, Melon EM 782 F 1 Enza Zaden Sembol, Sereen Golden Valley Seed Alia, Amur, GVS 125, GVS 205, GVS 206 Hazera Genetics Elario, Galante, Galapago, Jalisco, Lavi gal, Veronica, Vitorio, Nestor, Gal 52, Gal 152 Hyg rotech Omega Namdhari Seed NS 923, NS 929, NS 931 Nirit Seed Nirit, Gilat Nunhems Solarnet, Solarking, Esmeralda, Estoril, Malika, Solarprince Rogers/Syngenta Vicar, Galileo Seigers Seed (Seminis) Gallicum United Genetics Early Gal, Galistar, Green G o, Green Star, Rs Flavorite Technisem Caline Vigour Seeds NiZ 52 07 F1 Zeraim Gedera/Syngenta Arava, Don Juan, Inbar, Royal, Campeon, Jazmo, Pharis, Royal, Boa Vista,

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49 CHAPTER 3 PRODUCTION, EVALUATION, AND SELECTION OF ANTISENSE ACC OXIDASE ( CM ACO 1 ) GALIA F1 HYBRID MUSKMELON (Cucu mis melo L. var. reticulatus Ser. ) Introduction The Galia ( Cucumis melo L. var. r eticulatus Ser.) musk melon was developed in Israel by breeder Zvi Karchi and released in 1973. It was bred for production in the warm, dry d esert conditions in Israel (Karchi, 2000). The female parental line of Galia is a Ha Ogen type melon cultivar called Noy Yizreel (Karchi, personal comm., 2004). Noy Yizreel fruit are green -fleshed with a smooth, sutured skin (Figure 3 -1) (Karchi, 2000). Ha Ogen melons were introduced to Israel from Hungary and were a popular cultivar adapted to the intensive agricultural practices, which included the use of plasticulture with irrigation and fertilization (Karchi, 2000). The smooth skin of Ha O gen melon made it susceptible to damage and therefore, it was difficult to ship. Additionally, it was open -pollinated, which made it easy for others to grow and save seeds of these melons (Karchi, 2000). The male parental line of Galia was originally f rom the Peninsula of Crimea, in the Ukraine, and is a cultivar called Krymka, in which fruits were round with a golden, netted skin and light green, firm flesh (Karchi, personal comm., 2004). Krymka was selfed over 10 generations and selected for unifo rmity (Karchi, personal comm., 2004). The resulting hybrid cross the Galia muskmelon (Figure 3 2) had round fruits with an attractive orange -netted skin and a green, mellow textured flesh with a unique aroma and high soluble solids content (13 to 15 Brix) and was a high yielding cultivar (Karchi, 2000). In addition, Galia muskmelon was considered to have a longer shelf -life compared with the Ha Ogen types and therefore was able to compete in the European

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50 markets (Karchi, 2000). Furthermore, Galia muskmelon was an F1 hybrid, an exclusive, Israeli cultivar (Karchi, 2000). As a result of marketing campaigns with Agrexco and sales at the British food chain, Marks and Spencer, Galia muskmelons became popular all over Western Europe, except France (K archi, 2000). The popularity of this melon made it into a new market class of melons as breeders later released Galia -type (GT) cultivars with improved shelf life and disease resistance (Karchi, 2000). At present, there are over 75 GT cultivars available from numerous seed companies (Harty, 2009). Today Galia muskmelons have a reduced shelf life as compared with newer GT cultivars, which often lack the flavor and high SSC of the original Galia (Mitchell et al., 2007). In order to maintain the high quality and flavor of the original Galia muskmelon, the Galia male parental line (cv. Krymka) was transformed with an antisense ACC oxidase gene (CMACO 1) (Nuez -Palenius et al., 2006a ). ACC oxidase is the catalyst in the last step of the ethylene biosynthetic pathway (Yang and Hoffman, 1984). Ethylene is a hormone that has a major role in fruit ripening and senescence (Abeles et al., 1992), initiates fruit softening, changes in carbohydrate metabolism, a roma volatile production and abscission. Fru it size and sugar content are not regulated by ethylene (Pech et al., 1999; reviewed by Nuez -Palenius et al., 2008). The work of Nuez -Palenius et al. (2006a ) produced two independent antisense Krymka lines named TGM -AS 1 and TGM -AS 2. Fruits from TG M -AS 2 line produced less ethylene and were firmer than untransformed fruits at half and full -slip stages (Nuez -Palenius et al., 2006b ). TGM -AS 1 fruits also exhibited lower ethylene production, but only during the half -slip stage (Nuez Palenius et al. 2006b).

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51 Conversely, the female parental line of Galia muskmelon, cultivar Noy Yizreel was unable to be transformed with an antisense ACC -oxidase (CMACO 1) gene (Nuez Palenius et al., 2005). Nuez -Palenius developed antisense Galia muskmelon hyb rids (T0GMH -AS 1 and T0GMH -AS 2) produced from the T0 transgenic male parental lines (TGM -AS 1 and TGM -AS 2) (Mitchell et al., 2007b). During fall 2004, T0 TGMH -AS 1 and T0 TGMH -AS 2 lines remained on the vine five days longer than Galia (Mitchell et al ., 2007b). However, in a previous crop, a severe powdery mildew ( Podosphaera xanthii ) epidemic led to no differences in days to harvest (DTH) between antisense lines and Galia (Mitchell et al., 2007b). Due to these challenges, it was desirable to obtain AS Galia hybrids where both the male and female line incorporated the antisense ACC oxidase (CMACO 1) gene. With both parents positive for the transg ene, it was hypothesized that there would be an increased chance of the F1 hybrid progeny fruit to have reduced ethylene production and therefore a longer shelf -life. As an alternative to plant transformation, since that previously did not work in the female line (Nuez -Palenius et al., 2005) the backcross method was used to insert the antisense ACC oxid ase ( CM ACO 1) gene into the female line. Pollen from transgenic F1 Galia (TGH1 and TGH2) was used to pollinate the wild -type female Galia parental line. Female transgenic backcross 1 (BC1) seeds were produced during the summer of 2004. Repeated back crossing continued until the genetic background of the female was 97% (BC4). Backcrossing is a type of recurrent hybridization used to add a desirable allele to an already adapted and productive cultivar that is lacking in the desired allele (Poehlman and Sleper, 1995). Therefore, t he objective s of this research were to produce an elite stock of the TGM -AS 1 and TGM -AS 2 lines, use these lines to produce an

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52 antisense ACC -oxidase (CMACO 1) female parental line through backcrossing, produce an antisense Ga lia (ASG) hybrid muskmelon with both female and male transgenic parental lines through traditional breeding methods, and collect fruit quality data (fruit size, firmness, soluble solids content (SSC), ethylene and respiration) at different stages of growt h on preliminary lines for selection of ASG hybrids to be used for further research. Future research w ould include evaluation of fruits at different harvest stages for fruit quality, including size, SSC, firmness, and aroma volatiles. Ethylene and respir ation rates will be observed in order to verify reduced ethylene in transgenic lines as well as track the climacteric. These results will help establish fruit quality characteristics of ASG muskmelons and develop guidelines of when to harvest the ASG musk melons in order to benefit from the transgenic modification. Materials and Methods Antisense ACC-oxidase Galia F1 Hybrid D evelopment Development of elite parental lines, bearing an antisense ACC -oxidase gene was done throughout 2004 and 2005. TGM -AS 1 a nd TGM -AS 2 (T0 generation) were selfed and selected for the delayed ripening phenotype until a homozygous T4 generation was produced. At the same time, the T0 hybrid lines, TGH -AS 1 and TGH -AS 2 lines were crossed to the female parental line and an antise nse (AS) backcross 1 (AS BC1 generation) was also produced. The AS BC1 was used to continue to backcross the transgene into the female line until a backcross 4 (AS BC4) female population was produced. Parental line production was done in an evaporative -c ooled fan and pad glasshouse according to the methods of Nuez -Palenius et al. (2006a). Plants were grown using commercial production, pruning and nutrient requirements according to the methods of Shaw et al. (2001).

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53 Selfing and backcrossing was complete d each morning, during flowering, between 7:30 a.m. and 11:00 a.m. To self, three male flowers were removed from the plant and all three flowers were used to pollinate one female flower on the same plant. To backcross, three male flowers were picked from a designated backcross plant. These three flowers were then used to pollinate one female flower on a wild -type female plant. Prior to the backcross pollination, the female flower was emasculated with sterile tweezers. All pollinated flowers were tagged with the date Two crops of selfed males and backcrossed females were able to be produced each year, one from January to May and again from August to October. Production continued until AS male T4 and AS female BC4 populations were produced in fall 2 005. Data w ere recorded during each season for days to harvest (DTH) for the male transgenic lines. Every fruit from a plant was given a reference number. The fruits with the longest DTH were selected for the next generation. Also, during fall 2005, T3 TGM -AS 1 and TGM -AS 2 male lines and a wild type (WT) male were screened for ethylene and respiration data. This was done to observe both the amount of ethylene evolved from the new lines and track the climacteric peaks in these lines, which would provid e a new method of when to harvest the antisense hybrids. Fruits were harvested at different days after anthesis. Ethylene and respiration were collected according to the procedures as outlined for the hybrid evaluation. During spring 2006, the female A S BC4 and AS T4 male were crossed and AS Galia (ASG) hybrid seeds were produced. The ASG hybrids were made in different combinations (AS

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54 two independent transgenic male lines from Nuez -Palenius et al. (2006a ), these two lines resulted in two main ASG hybrid family lines, ASG 1 and ASG 2. Transgene Detection for Antisense Male, Female and Hybrid Lines All seedlings were produced at the University of Florida, Gainesville, FL campus in a Conviron plant growth chamber (Controlled Env. Ltd.,Winnipeg, Manitoba, Canada) according to Mitchell Harty et al. ( 2009 a). DNA extraction was done when seedlings had one true leaf, according to Mitchell Harty et al. (200 9 b). A polymerase chain reaction (PCR) analysis was used to identify the seedlings with the transgene. The PCR reaction was conducted in a DNA Therma l Cycler 480 (Applied Biosystems, Foster City, CA U.S.A. ) according to the parameters of Nuez -Palenius et al. ( 2006a) Following amplification, the PCR products were viewed on a 1% agarose gel by ultraviolet (UV) light according to Nuez Palenius et al (2006a ). PCR analysis was completed on every putative transgenic seedling prior to planting. ASG M uskmelon Production Two ASG muskmelon trials were conducted, in fall 2006 and spring 2007. The first trial, completed in fall 2006, involved the screening of multiple lines of the ASG material. Seeds were sown on 7 July 2006. The commercial Galia and ASG lines used were Galia muskmelon (Hazera Genetics, Israel ), and three lines each of ASG 1 hybrid combinations of ASxWT (ASxWT(ASG 1a), ASxWT(ASG 1b) and ASxWT(ASG 1c)), WTxAS (WTxAS(ASG 1d), WTxAS (ASG 1e) and WTxAS (ASG 1f)) and ASxAS (ASxAS(ASG 1g), ASxAS(ASG 1h) and ASxAS(ASG 1i)), and one ASxAS line of ASG 2, labeled ASxAS2 was also sown. After the fall 2006 trial, one ASG line from each cross (ASxWT, WTxAS and ASxAS) were selected and evaluated with the original

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55 Galia hybrid (Hazera Genetics, Israel) in spring 2007. In spring 2007, seeds were sown on 19 Jan. 2007. Once transg enic seedlings were identified through PCR analysis and all seedlings ha d three true leaves, they were transplanted on the 3 Aug. 2006 and 27 Feb. 2007. The plants were grown according to the production and nutrient requirements of Shaw et al. (2001) in a saw -tooth style, passively -ventilated greenhouse (TOP greenhouses, Ltd. Barkan, Israel), located at the University of Florida, Plant Science Research Education Unit located in Citra, FL. ASG hybrids were not self -pollinated. B umble bees from Class A research hives ( Bombus impatiens Natupol, Koppert Biological Systems, Inc ., Romulus, MI ) were used for cross -pollination. All flowers were tagged with the date of anthesis to track the days to harvest (DTH). An integrated pest management (IPM) approach, which used scouting, biological control and sprays, was used for manageme nt of arthropod pests according to Mitchell Harty et al. (200 9 a). Fruit Harvest Procedure Fruits were harvested from 29 Sept. to 30 Oct. 2006 and 10 May to 18 June 2007 according to the methods of Mitchell Harty et al. (200 9 a) at four stages of ripening Stage: 1.) zero -slip green (ZG): external skin still green in color with no abscission layer development; 2.) zero -slip, yellow -green (ZYG): external skin green and yellow, with no abscission layer development; 3.) half -slip (HS): fruit abscising half -w ay; and 4.) full -slip (FS): fruit separates easily from the stem. During each harvest period, fruits were picked each afternoon, transported to the Gainesville campus and stored at 20 C for 12 hours. All postharvest variables were measured the following morning, 12 hours after harvest.

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56 Ethylene, R espiration and Fruit Quality M easurements Ethylene and respiration measurements were measured on whole fruits consistent with methods and equipment as described by Mitchell Harty et al. (200 9 a). A fter the fr uits had their 12-hour storage interval (following harvest), ethylene and respiration rates were measured from all fruits. Following ethylene and respiration measurements, fruit quality variables, which included flesh thickness, firmness, and S SC were measured from a 2.5 cm slice of fresh pulp from the equatorial region of the fruit according to the methods of Mitchell et al. (2007a). Pulp firmness was determined at two equidistant points on the equatorial region of each fruit slice using the Instron Uni versal Testing Instrument (Model 4411C8009, Canton, MA), which was fitted with a 50 kg load cell and an 11 -mm convex pro be with a crosshead speed of 50 mm min1. SSC (Brix) was measured with a temperaturecompensating, handheld refractometer (Model 10430, Reichert Scientific Instrument, Buffalo, NY) from fresh juice expressed from two pulp samples; and flesh thickness was measured with a caliper (Digimatic Mycal, Mitutoyo, Japan) from peel to cavity. Statistical A nalysis The ASG hybrid trials were conducted in a randomized complete block design (RCBD) with four replications. Number of fruits per treatment (n) ranged from one to 10 fruits. However, in fall 2006, some of the ASG lines did not produce sufficient fruits to be harvested at stage ZG. Th eref ore fall 2006 ZG results are presented as grouped data (ASxWT (ASG 1a,b,c), WTxAS (ASG 1d,e,f) and ASxAS (ASG 1g,h,i)). For stages ZYG, HS and FS, results are presented by the individual lines After promising individual ASG lines were selected from the fall 2006 results, the selections were evaluated again during spring 2007. Fall and spring results from the selections were then

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57 analyzed together at each stage. All data were subject to analysis of variance (ANOVA) using the GLM procedure (SAS Institut e, Version 9, Cary, NC, U.S.A.). Significant means were separated with Fishers Least Significant Difference (P<0.05). Standard error (SE) values were calculated for each ethylene and respiration data point. Results and Discussion Parental Line P roduction Selfed AS male line TGM -AS 1 over the four generations, averaged an extra 22 days on the vine as compared with the wildtype male (Table 3 1 and Figure 33). DTH for T2 to T4 generations of TGM -AS 2 decreased over the four generations, which indicated tha t the delayed -ripening phenotype was not expressed. The progressed failure of line TGM -AS 2 is interesting, since this line had previously demonstrated the greatest inhibition of ethylene production in the original T0 work by Nuez Palenius et al., (2006b ). Review of the southern blot analysis by Nuez Palenius et al. (2006a) illustrated a single copy of the transgene for both TGM -AS 1 and TGM -AS -2 lines. However, the photograph (Nuez -Palenius et al., 2006a) portrays line TGM -AS 2 with a higher expres sion of the transgene as compared with line TGM -AS 1. Possibly, multiple insertions of the transgene occurred which could lead to gene silencing (Vaucheret and Fagard, 2001). This could be a possible explanation for the failure of this line to express th e desired reduced ethylene delayed ripening characteristic. Backcross 4 (BC4) female lines with the antisense ACC -oxidase (CMACO -1) were produced from both TGM -AS 1 and TGM -AS 2 male parental lines. AS BC female lines were selected based on transgene dete ction only. Comparison of the antisense (AS) BC female lines to a wild -type female was not easily measured by DTH. Wild type fruit from female line plants averaged 47 to 57 DTH, as did DTH for fruits

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58 from AS BC female lines (Table 3 2). Production of f emale backcross lines was also difficult as compared with selfed male production, as many female line backcrossed flowers did not set. This may have been due to damage to the ovary while being emasculated. Ethylene and respiration rates were to be analyz ed from T3 TGM -AS 1, TGM AS 2 and wild-type males at different days after anthesis. However, since these days varied for each fruit, as did the number of fruit collected for each day, results were presented according to different stages of ripening stage ZG: zero -slip with green skin ; stage ZYG: zero -slip with yellow/green skin; stage HS: half -slip; stage FS: full-slip; and stage PS: 1 day after full -slip, or post -slip. Ethylene and respiration from T3 and wild type (WT) fruit from male line plants gener ally increased during stage ZYG (Figs. 3 4 to 3 6). TGM -AS 1 fruits had low ethylene production at stages ZG, ZYG and HS, which was followed by increases at FS and PS (Fig. 3 4). Compared to fruit from WT male lines, TGM -AS 1 had lower ethylene productio n at all stages except stage PS. TGM -AS 2 male fruits had diverse amounts of ethylene production (Fig. 3 5), which were similar to Galia at stages ZYG and FS. Due to the continued decline in delayed ripening for TGM -AS 2 fruits and the varied ethylene production at all stages, only an ASxAS hybrid combination of line TGM -AS 2 was considered useful for subsequent screening. The instability of the T3 TGM -AS 2 line is interesting, considering that the T0 TGM -AS 2 line from Nuez -Palenius et al. (2006b) p roduced the least amount of ethylene. Hybrid M uskm elon R esults and Selection, Fall 2006 Although there was some variation among the lines in DTH during every stage, there was a general pattern in the ASG 1 lines in which they had with the greatest DTH, while line ASG 2 and Galia had the lowest DTH. At stage ZG (Table 3 3), no

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59 significant differences in DTH occurred among the lines. Fruit quality at stage ZG illustrated a difference in only SSC among the lines (Table 3 3). The ASG 1 lines, ASxWT and ASxAS had the lowest SSC. Stage ZG muskmelons from all lines weighed greater than 1 kg and were firm with low ethylene and respiration rates (Table 3 3). At stage ZYG (Table 3 4), DTH was greatest for ASG 1 lines, ASxWT (c), and ASxAS (h,i) while Gali a had the least DTH. Line ASxAS2 (ASG 2) was similar in DTH to Galia. Stage ZYG fruit exhibited the most differences among lines in fruit quality (Table 3 4). Mean fruit weight for all lines was again greater than 1 kg. All lines, except ASxWT (ASG 1 d) and ASxAS (ASG 1i), had SSC above 9 Brix, deeming them acceptable to meet the standards for U.S. Grade No.1 fruit (Lester and Shellie, 2004). Fruits were generally still firm at this stage; however the ASG 1 lines, ASxAS (g,h,i) and ASxWT (b) were fir mer than Galia. Lines WTxAS (ASG 1d) and ASxAS2 (ASG 2) were similar in firmness to Galia. Ethylene production rates were lowest for the ASG 1 lines, ASxWT (b,c) and ASxAS (h,i) and highest for Galia and lines WTxAS (ASG 1d) and ASxAS2 (ASG 2). The similarities in quality and ethylene production between Galia and line ASxAS (ASG 2) continued to verify that the transgene in line ASG 2 has probably been silenced. At stage HS, DTH was greatest for the ASG 1 lines, ASxAS (i) and WTxAS (g) (Table 3 5 ). Fruit weight and size varied among the lines, however, all lines weighed greater than 1.2 kg (Table 3 5). Fruit SSC was at or above 9 Brix for all lines. Ethylene and respiration rates increased for all lines from stage ZYG (Table 3 5). For fruits h arvested at stage FS, all ASxAS (ASG 1g,h,i) lines remained on the vine longer than Galia (Table 3 6). Overall, the ASG 1 lines, ASxAS (g,h,i) remained

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60 on the vine an average of eight days longer than Galia. The maximum DTH for lines of ASxAS (ASG 1) ranged from 57 to 60 DTH 10 to 13 days longer than the max DTH for Galia. DTH for line ASxAS2 (ASG 2) was similar to Galia, which continued to confirm gene silencing in this line. Stage FS fruit exhibited few differences in fruit quality among all th e ASG lines and Galia (Table 3 6). Fruit SSC was again above 9 Brix for all lines, though many were at or above 11 Brix. Muskmelon SSC of 11 Brix or higher meet the criteria of U.S. Grade Fancy (Lester and Shellie, 2004). Fruit firmness and ethyl ene production rates varied among lines Galia was among the firmest fruits, though it also produced among the greatest amount of ethylene evolution. This was most likely due to the fruits being at or near their climacteric peak ( Lyons et al., 1962). L ine ASxAS (ASG 1i) was among the least firm fruit, and also had the lowest ethylene production. This was probably due to the longer time on the vine and failure of the ASxAS (ASG -1i) fruit to slip early during its peak climacteric phase. Delay in abscissi on zone development was also observed in T0 TGM -AS 2 muskmelons (Nuez -Palenius et al. (2006b) as well as in antisense ACO Charentais cantaloupes (Flores et al., 2001). Respiration rates were similar among all lines (Table 3 6). Since the ASG 1 lines, par ticularly those in lines ASxAS and ASxWT, had differences in DTH, fruit firmness and ethylene production at stage ZYG, this stage appeared to be the ideal time to harvest ASG 1 fruit as to obtain optimum quality and perhaps a longer shelf -life. At stage Z YG, all ASG 1 muskmelons had acceptable fruit weight and size as well as acceptable SSC (9 Brix). Fruit firmness for ASG 1 lines, ASxWT and ASxAS was also greatest and ethylene production was low. If ASG 1 fruits

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61 are not harvested at stage ZYG, then by stages HS and FS, they are equal in fruit quality (especially firmness) to Galia and present no added postharvest benefits. Therefore, the antisense Galia hybrid fruits could be harvested early (prior to full -slip), shipped, and then consumed at the high fruit quality standards of wild -type Galia. Within line ASxAS (ASG 1 g,h,i) each of the three individual lines tested exhibited the qualities desired for a longer shelf life Galia, especially if harvested at stage ZYG (Table 3 4). One of these l ines (ASG 1h) was selected for further research due to its high SSC (12 Brix) and low ethylene production (0.69 ng kg1 s1) at stage ZYG as well as averaging 46 DTH by stage FS. Within the ASxWT (ASG -1 b,c) lines, at stage ZYG, two of the lines tested e xhibited lower ethylene production than Galia and also had a high SSC, but only line, ASG 1b, was significantly firmer than Galia (Table 4). Fruits from line WTxAS (ASG 1 d,e) did not demonstrate many significant differences from Galia in terms of f irmness and ethylene production. The WTxAS line, perhaps behaved similar to the first T0 antisense hybrids grown in 2004 (Mitchell et al., 2007b), where only the male parental line incorporated the transgene. However, in order to confirm that line WTxAS (ASG 1 d,e) is not expressing the desired characteristics, a selection will be taken from this line and it will be included in future studies for additional analysis. The ASG 2 line, ASxAS2 displayed characteristics of Galia as it was similar in firmne ss and in ethylene production to Galia at most stages. Since this line stopped expressing the desired characteristics, was not used in subsequent ASG research. Only ASG 1 lines will be included in future research.

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62 The ASG 1 lines selected and produced in spring 2007 were ASxAS (ASG -1h), ASxWT (ASG 1b) and WTxAS (ASG 1d). These selections were also used to compare fall 2006 and spring 2007 results. Fall 2006 and S pring 2007 ASG -1 Results Stage ZG At stage ZG, only flesh thickness and SSC differed among lines, however there were seasonal differences for most variables. Spring 2007 fruits were generally smaller and lower in SSC, averaging 8 Brix (Table 3 7). There were no line x season interactions for any stage ZG variables over fall 2006 and spring 2007. This was probably due to the few differences observed am ong the lines, as all fruits were immature at this stage. Stage ZYG At stage ZYG, DTH differences occurred among the lines, where all ASG 1 lines were harvested later than Galia. Also, the ASG 1 lines were overall 50% firmer than Galia. Seasonal differences were also observed for every variable. Spring 2007 fruits were smaller and 60% less firm than fall 2006 fruits (Table 3 8). There were also line x season interactions for SSC, ethylene a nd respiration (Table 3 9). Fruit SSC for line WTxAS was not affected by season, whereas all other lines were sweetest in the fall. Fall melon crops have the potential of having higher brix value s because of low er night temperatures than spring crops (Le ster et al., 2007; Lester et al., 2006; Beaulieu et al., 2003). Both ethylene and respiration rates were increased from stage ZG fruit. Ethylene rates also varied between fall and spring, while respiration rates remained consistent, both among lines and seasons. The variation in ethylene rates might be due to an atypical series of wildfires that occurred during spring 2007 in north Florida. The occurrence of

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63 smoke, which consists of ethylene (Rodriguez, 1932), is known to induce ripening and reduce chlo rophyll in fruits and leaves (Kader, 2002). During the week of the first harvest, smoke from wildfires was in close proximity to the production site (Fig. 3 7), this could have led to some of the ASG 1 fruits ripening more rapidly, and also to some plant damage that was later observed on the muskmelon leaves. In spring 2007, ASG 1 and Galia fruits had similar ethylene rates at stage ZYG, though in fall 2006, lines ASxWT and ASxAS produced less ethylene than Galia. Respiration rates were generally sim ilar among all lines in both seasons. Stage HS At stage HS, all ASG 1 lines were harvested later than Galia. ASG 1 lines, ASxWT and ASxAS had the firmest fruits. While the ASG 1lines, ASxWT and ASxAS were firmer than Galia, the TGM -AS 1 half -slip fru its in the work reported by Nuez Palenius et al. (2006b) were not firmer than the wild type. Thus, perhaps the addition of the antisense female parental line in the ASG 1 hybrids has aided in the improvement of the antisense hybrid. The fall 2006 fruits were again, larger and firmer compared to spring 2007 (Table 3 10), which again alluded to the poor environmental conditions that occurred in spring 2007. Line x season interactions for length, SSC and ethylene occurred (Table 3 11). Ethylene rates were either increased or remained at levels similar to stage ZYG. Fall 2006 ethylene rates at stage HS were higher than spring 2007 rates for all lines except Galia. There were no differences in ethylene among all lines. Stage FS Galia and ASG 1 fruit s at stage FS were similar in many quality variables, expect in size, where Galia muskmelons were larger and weighed more (Table 3 12). Stage FS fruit firmness decreased for all lines from stage HS. Ethylene and respiration

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6 4 rates were increased from st age HS, yet there were no differences among the lines. Fruit quality in fall 2006 was generally better as compared with spring 2007. Fruits were larger in fall 2006 and SSC and firmness were also higher as compared with spring 2007 (Table 3 12). The onl y line x season interaction was for DTH, where during spring 2007, Galia averaged the same DTH as all ASG 1 lines. However, in fall 2006, all ASG 1 lines had longer DTH than Galia, ranging from three to seven more days on the vine (Table 3 13). Antise nse ACO Charentais cantaloupes also displayed a delay in slipping from the vine (Ayub et al., 1996; Flores et al., 2001). The spring 2007 result of Galias similarity in DTH among the ASG 1 lines could be due to the effects of the wildfire/smoke event. Th e damaged plants may have caused Galia to ripen slower in this case, as ethylene and respiration rates of Galia and ASG 1 lines were also similar. Ethylene rates were higher in fall 2006 at stage FS than spring 2007. Again, the fact that fruits remai ned on the plant longer in the spring as compared with fall may have attributed to the lower ethylene rates at stage FS. By the time fruits reached stage FS in spring 2007, they were most likely post -climacteric and produced less ethylene as compared to fa ll 2006 fruits, which ripened without any plant stress. ASG 1 muskmelons harvested at stages ZG, HS and FS exhibited many similarities to the original Galia muskmelon, especially in firmness and ethylene production. Only stage ZYG ASG 1 muskmelons from lines ASxAS and ASxWT demonstrated increased firmness and lower ethylene, but only when grown in optimal conditions, such as good temperatures (Min: 18 C to Max: 35 C), minimal disease/insect pressure, and no wildfires or hurricanes. There was also no difference in respiration among the ASG 1 lines and Galia at stage ZYG, or at any other stage. Other

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65 reports of antisense ACO Charentais cantaloupes report a lack of a climacteric rise in the antisense fruit compared with wild type fruit (Bower et al., 2002). Perhaps both reduced ethylene and respiration may be observed in the ASG 1 muskmelons during a storage treatment where the gas emissions can be tracked over time. The ASG 1 lines of ASxAS and ASxWT demonstrated the most potential for a longer shelf -life Galia muskmelon if harvested at stage ZYG and produced in an optimal environment (such as acceptable temperatures and low insect/disease pressures ) These ASG 1 lines exhibited similar fruit size (greater than 1 kg), similar SSC (ranged from 10.5 Brix to over 12 Brix) to original Galia, yet were firmer than Galia at stage ZYG. The ASG 1 lines, ASxAS and ASxWT also produced less ethylene than Galia during fall 2006. Even though reduced ethylene was not observed again in spring 2007, most l ikely due to the environmental problems associated with that season, the ASG 1 lines ASxWT and ASxAS were again firmer than Galia in spring 2007 and had a later DTH than Galia. These results are similar to what Nuez-Palenius et al. (2006a and 2006b), Ayub et al. (1996) and Guis et al. (1997) reported in other antisense acc -oxidase melons in terms of firmer fruit, low ethylene and similar fruit quality (fruit size and SSC). Summary Renowned for their flavor and sweetness, Galia ( Cucumis melo L. var. r eticulatus Ser.) melons have a reduced shelf life as compared with standard melons. The objective of this research was to develop a true Galia F1 hybrid with a longer shelf -life while maintaining sweetness. The Galia male parental line was previously transformed with an antisense ACC -oxidase gene (CMACO 1),which produced two independent T0, TGM AS 1 and TGM -AS 2, transgenic lines (Nuez -Palenius et al., 2006a). These lines were used in a traditional plant breeding program to produce antisense (AS) Ga lia F1 hybrids

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66 (AS Galia F1 (ASG) muskmelon crosses were grown and evaluated with the original Galia cultivar. After the fall 2006 trial, one ASG line from each cross (ASxW T, WTxAS and ASxAS) was selected and evaluated again with Galia in spring 2007. Fruits were harvested at four stages of growth: stage 1.) zero -slip, green (ZG); 2.) zero -slip, yellow green (ZYG); 3.) half -slip (HS); and 4.) full -slip (FS). Data were rec orded for days to harvest, fruit weight, size, flesh thickness, soluble solids content (SSC), firmness, ethylene and respiration (CO2). At stage ZG, all antisense (AS) muskmelons were similar in size, quality, ethylene and CO2 production to Galia. At sta ge ZYG, ASxAS and ASxWT melons were significantly firmer than Galia in fall and spring, yet only produced less ethylene than Galia in fall 2006. Also at stage ZYG, there were no differences in SSC among all lines during each season. ASxAS, WTxAS, ASxW T melons remained on the vine an average of three to five days longer than Galia and were similar in quality, ethylene and CO2 to Galia at the HS and FS stages. The differences that occurred in lines ASxAS and ASxWT at stage ZYG indicated a potential that a longer shelf life Galia muskmelon were achieved when harvested at stage ZYG.

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67 Table 3 1. Days to harvest (DTH) results of the transformed (T) antisense mal e (TGM AS 1 and TGM -AS 2) lines that were selfed from spring 2004 through fall 2005. Days to harvest (DTH) Line T 1 T 2 T 3 T 4 TGM AS 1 51 52 59 59 TGM AS 2 4 6 4 2 37 36 Wild type 30 38 30 35 LSD (0.05) z 8.4 10.9 2.9 4.6 z Mean separation by Fishers least significant difference test (P Table 3 2 Days to harvest (DTH) results of backcrossed (BC) antisense fem ale (TGF AS 1 and TGF -AS 2 ) lines from spring 2004 through fall 2005. Days to harvest (DTH) Line BC 1 BC 2 B C 3 BC 4 TGF AS 1 51 50 51 48 TGF AS 2 53 54 49 44 Wild type 57 51 51 47 LSD (0.05) z 3.2 1.9 z Mean separation by Fishers least significant difference test (P

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68 Table 3 3 Stage ZG means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and grouped Antisense Galia (ASG 1 and 2) muskmelons, fall 2006. Line Days to harvest (DTH) Weight (kg) Length (mm) Width (mm) Flesh t hickness (mm) Soluble solids c ontent (Brix) Firmness (N) Ethylene (ng kg 1 s 1 ) Res piration (g CO 2 kg 1 s 1 ) Galia 38 1.53 155 143 35.9 10.5 39.8 0.21 9.2 ASxWT (ASG 1) 40 1.09 138 129 28.5 9.9 43.9 0.09 8.2 WTxAS (ASG 1) 39 1.40 153 141 30.7 6.3 37.3 0.22 8.0 ASxAS (ASG 1) 43 1.12 137 129 30.9 7.6 43.7 0.10 6.9 ASxAS (ASG 2) 40 1 .18 138 129 31.1 10.1 37.7 1.40 10.0 LSD (0.05) z 2.0 z Mean separation by Fishers least significant difference test ( P 0.05).

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69 Table 3 4 Stage ZYG means of days to harvest, fruit quality, ethylene production and respiration rates of Gali a, and individual lines of Antisense Galia (ASG 1 and 2) muskmelons fall 2006. Line Days to harvest Weight (kg) Length (mm) Width (mm) Flesh t hickness (mm) Soluble solids c ontent (Brix) Firmness (N) Ethylene (ng kg 1 s 1 ) Respiration (g CO 2 kg 1 s 1 ) Galia 38 1.50 151 143 30.5 11.8 24.9 2.86 10.9 ASxWT (ASG 1b) 41 1.30 145 136 31.2 12.2 38.1 1.41 11.7 ASxWT (ASG 1c) 43 1.67 142 138 30.3 11.2 28.0 1.40 13.4 WTxAS (ASG 1d) 42 1.42 149 140 30.5 9.5 28.0 2.52 16.0 ASxAS (ASG 1g) 41 1.45 149 145 30.4 8.5 32.8 1.97 15.8 ASxAS (ASG 1h) 43 1.30 144 134 30.5 11.6 33.4 0.69 10.4 ASxAS (ASG 1i) 45 1.31 156 137 32.9 6.75 31.3 0.69 8.8 ASxAS2 (ASG2) 40 1.60 157 144 35.8 12.0 21.7 1.95 13.4 LSD (0.05) z 3.5 1.8 6.4 1. 2 z Mean separation by Fisher s least significant difference test (P

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70 Table 3 5. Stage HS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia, Galia type (MG10183) and Antisense Galia (ASG 1 and 2) muskmelons, fall 2006. Line Day s to harvest Weight (kg) Length (mm) Width (mm) Flesh t hickness (mm) Soluble solids c ontent (Brix) Firmness (N) Ethylene (ng kg1 s1) Respiration (g CO 2 kg1 s1) Galia 40 1.43 143 141 34.2 12.3 21.4 2.84 12.1 ASxWT (ASG 1b) 43 1.80 162 148 36.4 12. 3 24.3 3.53 12.7 ASxWT (ASG 1c) 42 1.29 143 137 26.5 10.0 9.1 4.21 14.9 WTxAS (ASG 1d) 42 1.75 162 149 31.9 10.0 15.3 5.66 12.7 WTxAS (ASG 1e) 45 1.49 155 142 31.6 10.4 20.3 2.99 24.7 ASxAS (ASG 1g) 41 1.48 157 140 33.8 9.6 17.7 3.93 10.7 ASxAS (ASG 1 h) 43 1.45 147 140 32.8 11.8 25.6 2.37 12.3 ASxAS (ASG 1i) 51 2.05 166 157 36.8 9.0 11.2 1.79 14.9 ASxAS2 (ASG2) 37 1.72 163 146 33.8 11.0 18.0 6.93 12.7 LSD (0.05) z 4.6 0.4 12.9 1.9 7.8 2.3 z Mean separation by Fishers least significant differen ce test (P

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71 Table 3 6. Stage FS means of days to harvest, fruit quality, ethylene production an d respiration rates of Galia and Antisense Galia (ASG 1 and 2) muskmelons, fall 2006. Line Days to harvest DTH Min DTH Max Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Soluble solids c ontent (Brix) Firmness (N) Ethylene (ng kg1 s1) Respiration (g CO2 kg1 s1) Galia 39 32 46 1.68 157 147 33.2 11.3 18.1 4.97 14.0 ASxWT (ASG 1b) 43 34 53 1.36 145 136 33.2 12.1 18.9 5.07 13.7 ASxWT (ASG 1c) 42 37 47 1.12 141 131 29.2 9.9 12.3 3.08 13.2 WTxAS (ASG 1d) 43 38 49 1.31 145 133 29.4 10.3 13.1 5.83 15.0 WTxAS (ASG 1e) 41 37 47 1.41 155 140 30.4 11.1 16.4 4.34 14.6 ASxAS (ASG 1g) 47 37 58 1.86 165 154 37.8 9.2 14.2 3.36 11.8 ASxAS (ASG 1h) 46 40 57 1.45 150 137 34.2 11.1 16.2 3.03 11.8 ASxAS (ASG 1i) 47 43 60 1.54 157 142 37.3 9.5 9.7 1.13 10.7 ASxAS2 (ASG2) 40 34 47 1.60 157 144 36.3 12.1 13.0 5.55 12.0 LSD (0.05) z 5.3 1.8 4.7 1.8 z Mean separation by Fishers least significant difference test (P

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72 Table 3 7. Stage ZG means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antisense Galia (ASG 1) muskmelons, fall 2006 and spring 2007. Line Days to Harvest Weight (kg) Length (mm) Width (mm) Fles h thickness (mm) Soluble solids c ontent (Brix) Firmness (N) Ethylene (ng kg1 s1) Respiration (g CO 2 kg1 s1) Galia 47 1.34 148 134 32.7 9.6 36.5 0.13 6.91 ASWT 46 1.03 135 123 26.3 8.8 41.8 0.07 6.26 WTAS 49 1.27 146 132 29.5 7.1 37.6 0.11 5.50 ASAS 51 1.26 139 130 28.8 9.0 41.6 0.04 5.50 LSD (0.05) z 3.8 1.6 Fall 2006 40 1.34 145 136 30.9 9.4 40.8 0.15 7.89 Spring 2007 57 1.11 139 124 27.8 7.8 37.9 0.02 4.19 Significancey ** ** ** ** ** ** NS ** z Mean separation by Fishers lea st significant difference test (P y NS, *, ** Non -significant (NS) or significant F test at P

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73 Table 3 8 Stage Z Y G means of days to harvest, fruit quality, ethylene production and respiration rates of Galia a nd Antisense Galia (ASG 1 ) muskmelons, fall 2006 and spring 2007. Line Days to Harvest Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Firmness (N) Galia 41 1.24 142 132 29.0 17.4 ASWT 45 1.13 137 126 28.7 32.5 WTAS 46 1.08 133 127 29.2 2 1.2 ASAS 45 1.08 135 125 27.7 28.8 LSD (0.05) z 2.3 3.3 Fall 2006 41 1.38 147 139 30.8 30.7 Spring 2007 47 0.89 127 117 26.5 19.2 Significance y ** ** ** ** ** z Mean separation by Fishers least significant difference test (P y *, **, significant F -test at P respectively. Table 3 9. Stage ZYG line x season (L x S) interaction means of soluble solids content, ethylene production and respiration rates Soluble solids content (Brix) Ethylene (ng kg 1 s 1 ) Respiration (g CO 2 kg 1 s 1 ) Line Fa06 Sp07 Fa06 Sp07 Fa06 Sp07 Galia 11.8 9.8 2.86 1.07 10.89 9.64 ASWT 12.2 8.9 1.41 1.38 11.71 10.94 WTAS 9.2 9.4 2.75 1.89 16.99 8.55 ASAS 11.6 8.7 0.68 1.18 10.34 10.29 LxS LSD (0.05) z 0.95 0.74 2.9 z Mean separation for line x season interaction by Fishers least significant difference test (P

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74 Table 3 10. Stage HS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and A ntisense Galia (ASG 1) muskmelons, fall 2006 and spring 2007. Line Days to Harvest Weight (kg) Width (mm) Flesh thickness (mm) Firmness (N) Respiration (g CO 2 kg1 s1) Galia 42 1.21 130 31.9 16.2 12.20 ASWT 46 1.39 133 31.4 20.7 11.43 WTAS 49 1.28 132 30.2 14.6 13.67 ASAS 45 1.14 127 28.7 20.9 11.81 LSD (0.05) z 2.1 4.0 Fall 2006 42 1.56 142 33.8 22.1 13.50 Spri ng 2007 47 0.94 118 27.2 14.1 11.05 Significance y ** ** ** ** ** z Mean separation by Fisher s least significant difference test (P y *, **, significant F test at P Table 3 11. Sta ge HS line x season (L x S) interaction means of fruit length, soluble s olids content and ethylene Length (mm) Soluble sol ids content (Brix) Ethylene (ng kg 1 s 1 ) Line Fa06 Sp07 Fa06 Sp07 Fa06 Sp07 Galia 143 134 12.3 9.8 2.84 2.48 ASWT 162 131 12.3 8.7 3.54 1.8 WTAS 157 133 9.3 8.9 3.89 1.65 ASAS 147 122 11.8 9.0 2.37 1.9 LxS LSD (0.05) z 7 1.1 0.7 z Mean separatio n for line x season interaction by Fishers least significant difference test (P

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75 Table 3 12. Stage FS means of days to harvest, fruit quality, ethylene production and respiration rates of Galia and Antisense Galia (ASG 1 ) muskmelons, fall 2006 and spring 2007. Line Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Soluble s olids c ontent (Brix) Firmness (N) Ethylene (ng kg1 s1) Respiration (g CO 2 kg1 s1) Galia 1.39 148 135 32.5 10.9 14.6 3.89 13.4 ASWT 1.12 136 126 30.5 10.9 16.6 4.03 13.6 WTAS 1.08 136 125 29.4 9.19 12.3 3.78 13.3 ASAS 1.15 137 126 30.1 10.1 13.6 3.01 12.0 LSD (0.05) z 0.2 6. 9 6.4 0.9 2.0 Fall 2006 1.46 151 139 33.1 11.2 17.0 4.38 13.6 Spring 2007 0.91 128 117 28.1 9.32 11.5 2.98 12.6 Significance y ** ** ** ** ** ** z Mean separation by Fishers least significant difference test (P y NS, *, ** Non -significant (NS) or significant F test at P

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76 Table 3 13. Stage FS means of significant line*season interaction means of days to harvest Days to harvest Line Fa06 Sp07 Gali a 39 46 ASWT 43 47 WTAS 42 47 ASAS 46 49 LxS LSD (0.05) z 1.5 z Mean separation for line x season interaction by Fishers least significant difference test (P Figure 3 1. Galia muskm elon parental lines. The female parental line, Noy Y izreel (Left) and the male parental line, Krymka (right). Figure 3 2. The Galia F1 hybrid muskmelon.

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77 TGMAS1 Wild type Figure 3 3. TGM -AS 1 fruit from a T4 generation still green a nd on the vine after pollination on 3 23 06; and a wild-type male fruit ready to be harvested at full -slip that was pollinated on 3 25 06. Figure 3 4. Ethylene evolution and respiration (CO2) rates of T3 TGM -AS 1 muskmelons harvested at d ifferent stages (ZG= zero -slip, green; ZYG= zer o -slip, yellow/green; HS= half -slip; FS= full -slip; PS= post -slip) o f ripening, fall 2005. 0 5 10 15 20 25 ZG ZYG HS FS PS Stage g CO2 kg-1 s-1 0 2 4 6 8 10 ng kg-1 s-1 CO2 Ethylene

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78 TGM2 0 5 10 15 20 25 ZG ZYG HS FS PS Stage g CO2 kg-1 s-1 0 2 4 6 8 10 ng kg-1 s-1 CO2 Ethylene Figure 3 5. Ethylene evolution and (CO2) respiration rates of T3 TGM -AS 2 muskmelons harvested at differen t stages (ZG= zero -slip, green; ZYG= zero -slip, yellow/green; HS= half -slip; FS= full -slip; PS= post -slip) of ripening, fall 2005. 0 5 10 15 20 25 ZG ZYG HS FS PS Stage g CO2 kg-1 s-1 0 2 4 6 8 10 ng kg-1 s-1 CO2 Ethylene Figure 3 6. Ethylene evolution and respiration (CO2) rates of wild -type male (Krymka) muskmelons harvested at different stages (ZG= zero -slip, green; ZYG= zero slip, yellow/green; HS= half -slip; FS= full -slip; PS= post -slip) of ripening, fall 2005.

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79 Figure 3 7. The Protected Agriculture Greenhouse site enveloped in smoke from nearby wildfires during the week of May 8, 2007.

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80 CHAPTER 4 GALIA MUSKMELON FRUIT QUALITY AND FLAVOR ( Cucumis melo L. var. reticulatus Ser. ) Introduction The Galia muskmelon features a golden -netted exterior, sweet, green flesh and a musky fragrance (Karchi, 2000) It is an F1 hybrid that was developed by Israeli breeder, Zvi Karchi, who named it after his daughter; it was released in 1973 (Karchi, 2000). After its introduction, Galia muskmelon quickly became a popular market name throughout Europe and the Medit erranean by way of an intense marketing campaign with Agrexco an Israeli agricultural exporter, and sales at the popular British food chain, Marks and Spencer (Karchi, 2000). By the 1980s, Galia muskmelons were distributed all over Western Europe, exce pt France (Karchi, 2000). Its popularity was attributed to its intense flavor, aroma and sweetness. In effect, Galia F1 hybrid melon production was one of the factors that helped revive Israels agricultural production, breeding and research as well a s increase its competive advantage in world markets (Karchi, 2000). Although Galia muskmelon is an exceptional fruit, it has some limitations. Besides being highly susceptible to powdery mildew ( Podosphaera xanthii) (Mitchell et al. 2006, 2007a and 2007b), another main disadvantage is its short shelf -life (Mitchell et al. 2007b; Nuez Palenius et al., 2006a and 2006b; Aharoni et al., 1993). In order to achieve peak flavor, Galia must be picked at a fully ripe or full -slip stage (Mitchell et al. 2 007b; Nuez Palenius et al., 2005), which reduces storage life. Exporters have increased shelf life, but decreased quality by harvesting fruit at an immature stage; however this leads to lower quality fruit (Fallik et al., 2001; Cantliffe and Shaw, 2002; Pratt. 1971).

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81 As a result of Galias limitations and its popularity, breeders have worked to improve disease resistance/tolerance as well as improve shelf -life (Mitchell et al., 2007a), resulting in a new generation of improved Galia muskmelons (MOA G, 2006). Today Galia is a trade name for other look a like melon cultivars, commonly c alled Galia type melons. T here are over 75 Galia type cultivars available in the market (Harty, 2008). Unfortunately, although these Galia type cultivars ar e firm, they often lack the flavor, aroma and high soluble solids content of the original Galia hybrid (Mitchell et al., 2007a and 2007b). This is also true for other crops, such as tomato (Solanum lycopersicum ), where breeders have improved traits such as yield and disease resistance, but have fallen short on improving fruit quality (Causse et al., 2002). Although flavor comprises taste, texture and aroma (Goff and Klee, 2006), it is a complex trait also determined by many factors including genetics, e nvironment, culture, production and postharvest handling (Baldwin, 2002). In order to improve taste, breeders must consider all this information to gain the desired quality (Causse et al., 2003). This includes consideration of the developmental and biochemical changes in fruit color, texture, flavor and aroma (Li et al., 2006). Sweetness, however, is often considered to be one of the most important fruit quality components in muskmelon (Yamaguchi et al., 1977). Nevertheless, since muskmelons tend to be quite variable in quality, aroma and sensory analyses are also used to determine muskmelon fruit quality (Beaulieu and Lea, 2003; Aulenbach and Worthington, 1974; Yamaguchi et al., 1977). Sensory measurements of quality attributes can provide an approxi mation of consum er acceptability (Abbott, 1999) as t aste is important aspect of organoleptic quality (Causse et al., 2002).

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82 The characteristic flavor that is associated with melons is dependent on its aroma. Aroma volatiles are released as the fruit rip ens and their presence, absence and quantity characterize each melon type (Pratt, 1971; Teranishi, 1971). According to Pratt (1971) research on muskmelon aroma began in the 1930s when Rakitan (1935, 1945) identified and noted increases during ripening of the compounds acetaldehyde and ethanol in muskmelons. Today, over 240 aroma compounds have been identified in muskmelon and the predominant aromatic compounds are esters, alcohols and aldehydes (Obando-Ulloa et al., 2008; Lamikanra, 2002; Beaulieu, 2006; Beaulieu and Grimm, 2001, 2003; Njissen et al., 1996). Aroma volatiles are believed to be under genetic control as there are marked differences between melon cultivars (Yahyaoui et al., 2002; Wyllie and Leach, 1992). Higher concentrations of aroma volati le compounds have been reported in mature cantaloupes (muskmelons) as compared with fruits at immature stages (Beaulieu, 2006; Senesi et al., 2005; Beaulieu and Grimm, 2001; Horvat and Senter, 1987). Aroma volatiles have been studied on various Galia typ e (GT) cultivars such as Arava, Fado, C8, 080, 7302 (Leach et al., 1989; Wyllie and Leach, 1992; Fallik, et al., 2001 and 2005; Hoberg et al., 2003; Obando-Ulloa et al., 2008; Shalit et al., 2001; Kourkoutas et al., 2006). To date, there are no known reports on the aroma volatiles of the original Galia muskmelon. The volatile work by Leach et al., 1989 and Wyllie and Leach (1992) may have been with the original Galia cultivar, but it was not stated; and the work by Kourkoutas et al. (2006) reported results on Galia muskmelons with no report of whether it was Galia or a GT.

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83 As it is already known and confirmed that Galia is a high -quality fruit with excellent flavor (Karchi, 2000; Shaw et al., 2001; Fallik et al., 2001; Aharoni et al., 1993), it is not known what distinguishes the original Galia muskmelon apart from some GT cultivars. To determine what may set Galia flavor apart from GTs, the present study focused on aroma, and sought to identify volatiles of the true Galia F1 hy brid. Materials and Methods Three experiments were conducted during fall 2006, spring 2007 and fall 2007. Seeds of Galia (Hazera Genetics, Israel) and MG10183 (Zeraim Gedera/Syngenta, Israel) were planted on 7 July 2006, 19 Jan. 2007 and 31 July 2007. An additional Galia -type (GT), Elario (Hazera Genetics, Israel) was only grown in fall 2007. Elario was added to the research experiment since it was identified as being one of the standard fall cultivars commonly grown in Israel (A. Gadiel, ARAVA, personal communication, 2007), whereas MG10183 has not yet been released (M. Peretz, Zeraim Gedera, personal communication, 2008). MG10183 was selected due to previous studies where it was found to be a higher quality (good SSC and firmness) GT mus kmelon (Mitchell et al., 2007a; Mitchell et al., 2006) Seedlings were produced at the University of Florida, Gainesville, FL campus in Model 128A polystyrene plug trays (Speedling, Bushnell, FL) with a commercial fine grade plug growing medium (Premier Pro Mix PGX, Quakertown, PA). Seedlings were grown in a Conviron plant growth chamber (Controlled Env. Ltd.,Winnipeg, Manitoba, Canada) at temperatures of 28 C (day) and 22 C (night) with 16 hour daily artificial lighting Seedlings were fertilized after the first true leaf expanded, and fertilization

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84 continued once per week with Peters Professional All Purpose Plant Food (Spectrum Group, St. Louis, MO) at the rate according to Mitchell et al. (2007). Once seedlings had three true leaves, they were tra nsplanted on the 3 Aug. 2006, 27 Feb. and 15 Aug 2007. Plants were grown in a saw tooth style, passively ventilated greenhouse (TOP greenhouses, Ltd., Barkan, Israel), located at the University of Florida, Plant Science Research Education Unit located in Citra, FL. The plants were grown using commercial greenhouse muskmelon production techniques and nutrient requirements according to the recommendations of Shaw et al. (2001). Plant spacing was 48 cm between plants and 90 cm between rows. Plant density was 2.5 plants m2. Pollination was achieved via bumble bees from Class A research hives ( Bombus impatiens Natupol, Koppert Biological Systems, Inc., Romulus, MI). In fall 2006 one hive was released on 23 Sept. 2006. In the spring there were three hives released on 29 March, 6 April and 26 April 2007; and in fall 2007 two hives were released on 29 Aug. and 20 Sept. 2007. All flowers were tagged with the date of anthesis. Temperature and photosynthetic photon flux (PPF) at the canopy level were recorde d daily at 30 -min. intervals by HOBO data loggers (Onset Comp. Corp., Bourne, MA). Within canopy temperatures were also recorded at 15 minute intervals by WatchDog data loggers (Spectrum Tech., Plainfield, IL). The monthly temperature averages were taken as an average of the within and at canopy readings. Insect pests were monitored weekly by scouting one plant from each plot per block. Biological control was used for management of arthropod pests in all three seasons. During all three seasons, white flies ( Bemisia tabaci biotype B), flower thrips (Frankliniella tritici (Fitch)), two -spotted spider mites ( tetranychus urticae ) and red

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85 mites (O ligonychus ilicis ) were observed in the crop. As a result, Erotmocerus eremicus and Encarsia formosa (ENERMIX Koppert Biological Systems, Inc., Romulus, MI) and Erotmocerus mundus (BEMIPAR, Koppert Biological Systems, Inc., Romulus, MI) were released to control whitefly at an average release rate of 7.4 wasps m2; Orius insidiosus (THRIPOR I, Koppert Biologica l Systems, Inc., Romulus, MI), a predatory bug for control of flower thrips was released at an average rate of 4.5 bugs m2; Amblyseius swirskii (SWIRSKI -MITE PLUS, Koppert Biological Systems, Inc., Romulus, MI), predatory mites of thrips and whitefly, w ere released at an average rate of 97 mites m2; Neoseiulus californicus (Biotactics Inc., Perris, CA) predatory mites, used to control two spotted spider mites were released at an average rate of 28 mites m2 and Neoseiulus fallacis (Biotactics Inc. Perris, CA) predatory mites, used to control red mites, were released at an average rate of 40 mites m2. Also released was a parasitic wasp, Aphidius colemani (APHIPAR, Koppert Biological Systems, Inc., Romulus, MI) of the green peach aphid ( Myzus pe rsicae [Sulzer]), as a preventative measure at an average rate of 3.3 wasps m2. Due to a spider mite infestation in certain areas of the crop during all three seasons, Abamectin miticide (Agri -Mek, Syngenta Crop Prot., Inc., Greensboro, NC) was sprayed (rate: 30 oz. ha1) on 26 Sept., 2006, 3 Oct., 2006 and 17 Sept. 2007. In spring 2007, an insecticidal soap, Mpede (Mycogen Corp., San Diego, CA) was sprayed to control spider mites (rate: 2% v/v solution). No preventative fungicides were sprayed for po wdery mildew ( Podosphaera xanthii ). Once powdery mildew was evident in the crop, plants were sprayed weekly with potassium bicarbonate (Milstop, BioWorks Inc., Fairport, NY; rate: 2.8 kg ha1), a foliar fungicide that suppresses powdery mildew and assists in

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86 keeping the plants productive. Milstop applications began on 3 Oct. 2006, 14 May 2007 and 8 Oct. 2007. Fruit Selection and Postharvest T reatments Fruits were harvested at full maturity, but at four different stages of ripening. Stage: 1.) zero -sli p green (ZG): external skin still green in color with no abscission layer development; 2.) zero -slip, yellow -green (ZYG): external skin green and yellow, with no abscission layer development; 3.) half -slip (HS): fruit abscising half -way; and 4.) full -slip (FS): fruit separates easily from the stem. Fruits were harvested from 29 Sept. to 30 Oct. 2006; 10 May to 18 June 2007 and 2 Oct. to 26 Oct. 2007. During each harvest period, fruits were harvested daily, in the afternoon and separated into two groups. I n the first group, the harvest treatment, all postharvest variables were measured 12 hours after harvest. In the second group, the storage treatment, fruits were stored at 20 C and 85% relative humidity (RH). Storage days varied for the different stage fruits. This was done to be able to track the climacteric phase (ethylene and respiration was measured daily from each fruit while in storage) and fruit quality data was measured at an appropriate edible time (not over ripe) Stages ZG and ZYG were store d for five days, stage HS was stored for three days, and stage FS fruit was stored for two days. Only fruits harvested during spring and fall 2007 were subjected to a storage treatment. The f all 2006 trial was used to determine proper aroma volatile coll ection methods at harvest only, a s torage treatment was not a part of that research. Immediately after harvest, fruit weight and size were recorded. Fruits were then transported to campus and put in 20 C storage. The next morning, 12 hours after harvest ethylene and respiration rates were measured from all fruits.

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87 Ethylene and Respiration M easurements To determine ethylene evolution and respiration rates, each melon was sealed in an airtight 3 L Tupperware container for one hour at 20 C allowing ethyl ene and carbon dioxide to accumulate. Two samples were taken from the headspace using a hypodermic syringe through a rubber septum. A 1.0 ml sample for ethylene was injected into a Tracor 540 Gas Chromatograph (Tremetrics Analytical Division, Austin, TX) equipped with a photoionization detector (PID) and a stainless steel alumina F1 column (Supelco, Sigma Aldrich, Bellefonte, PA), with a mesh size of 80/100 and was 914 x 3.18 mm (length x dia m .). The detector and injector operated at 100 C and the oven wa s 50 C. The carrier gas was helium with a flow rate of 40 ml min1. A 0.5 ml sample for carbon dioxide was injected into in a Gow -Mac, Series 580 gas chromatograph (GC) (Gow -Mac Instruments, Bridgewater, NJ, U.S.A.). The Gow -Mac was equipped with a the rmal conductivity detector (TCD) and a 80/100 mesh Porapak Q column ( Agilent Tech., Inc., Santa Clara, CA) that was 1219 x 3.18 mm (length x diam.). The carrier gas was helium at a flow rate of 30 ml min1. The detector and injector operate d under ambient conditions (26 to 27 C) and the oven was at 40 C After ethylene and respiration measurements, fruits from the harvest treatment were removed from storage. Fruits in the storage treatment remained at 20 C. Fruit Quality M easurements Directly follo wing storage for all treatments, fruit quality variables which included flesh thickness, firmness and SSC were measured on fresh fruit according to Mitchell et al. (2007). A 2.5 cm slice was taken from the equatorial region of each fruit and flesh thick ness, firmness, and SSC were measured. A caliper (Digimatic Mycal, Mitutoyo, Japan) was used to measure flesh thickness from peel to cavity. Pulp firmness

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88 was determined at two equidistant points on the equatorial region of each fruit slice using an Instro n Universal Testing Instrument (Model 4411C8009, Canton, MA). The Instron was fitted with a 50 kg load cell and an 11 mm convex probe with a crosshead speed of 50 mm min1. Firmness data were expressed as the maximum force (bioyield point, in Newton) atta ined during deformation. Soluble solids content (SSC) (Brix) was measured with a temperature -compensating, handheld refractometer (Model 10430, Reichert Scientific Instrument, Buffalo, NY) from fresh juice expressed from two pulp samples taken from the eq uatorial slice. Remaining pulp was used for aroma volatile collection and a quantity was frozen for Total Titratable Acidity (T T A) an d pH measurements. Titratable Acidity (TT A) an d pH were measured with a 719 S Titrino (Metrohm Ltd., Hersisau, Switz.) fr om mesocarp flesh that was macerated, centrifu ged (Beckman, Model J2 21) for 15 min. at 15, 000 x g and the supernatant was filtered through cheese clothe. T T A was determined by pH titration to 8.1 with 0.1 N NaOH and calculated as percent malic acid equi valents. Aroma Volatile C ollection and Analysis Aroma volatiles were collected from 100 g of fresh, chopped mesocarp flesh (2 cm (L) x 1 cm (w)) from each fruit. Volatile collection was done according to Schmelze et al. (2003) with nonyl acetate as an in ternal standard. Fruit was inserted into Simex glass tubes (28 x 1.5 x 610 mm; Pegasus Glass). With t he aid of a vacuum pump, air, filtered through a hydrocarbon trap (Agilent Technologies, Palo Alto, CA), flowed through the tubes for 1 hour at 618 ml min1. Volatiles were collected on a Super Q column (30 mg Altech resin) and eluted with methylene chloride. Volatiles were separated on an Agilent Technologies DB 5 column (length x diam.: 30 x 0.25 mm) and

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89 analyzed on an Agilent Technologies 6890N (7683 Series) Gas Chromatograph (GC) (Agilent Tech., Inc., Santa Clara, CA). The GC was equipped with a flame ionization detector (FID) with a detection temperature of 280 C. The GC/FID had an inlet temperature of 220 C and an oven ramp from 40 to 250 C at 5 per minute; the carrier gas was helium. Retention times were compared with known standards and quantified with Agilent ChemStation software. Volatile peak identities were confirmed by an Agilent Technologies 5975 Gas Chromatograph/Mass Spectrometer (G C/MS). Thirty -eight volatile compounds were identified. Of the 38 compounds, 11 to 19 compounds, depending on cultivar and harvest stage, were considered to be significant contributors (SCs) to the overall aroma of Galia and both GTs. Total SCs for Galia by stage FS were 1 8 while MG10183 and Elario had 17 and 1 9 SCs, respectively (Table 4 1). Significant contributors to aroma were identified as a result of dividing the concentration of the compound (determined with GC/FID) by its known odor t hreshold value (OTV), resulting in the odor value (OV) of the compound (Bauchot et al., 1998; Teranishi et al., 1991). Compounds with OVs greater than one were considered significant contributors to the aroma. Odor threshold values (OTVs) were obtained i n the literature The SC compounds for all cultivars consisted of over 90% of the total identified volatiles (TIV) in every season at harvest and after storage, with the exception of fall 2006 harvest volatiles, where the SC compounds consisted of only 78% of the TIV. Sensory Evaluation A sensory analysis was done in spring 2008 with cultivars Galia, MG10183, Elario and an additional cultivar, Red Moon was included as a control. The same production practices were used in spring 2008. The Red M oon cultivar is a type of

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90 melon called European netted cantaloupe. These are characterized by their deep, orange/magenta pigmented flesh and a sutured/netted exterior. The R ed Moon is currently trademarked the Perfect Melon, boasting a guaranteed high SSC of up to 14 Brix (Mitchell Harty et al., 2008). For the sensory evaluation, all fruits were harvested at their recommended harvest stage. Cultivars Galia, MG10183 and Elario were harvested at stage FS. Red Moon was harvested at its recommended stagejust when a crack at the abscission layer (or slip) begins to develop (Mitchell Harty et al., 2008). SSC and firmness values were also obtained. The evaluat ion was performed at the University of Florida Food Science and Human Nutrition taste panel laboratory. Panelists consisted of random students, employees and visitors to the UF campus. The day before the taste panel, fruits were harvested at stage FS for cultivars Galia, MG10183 and Elario. Red Moon was harvested at the onset of abscission layer development at the stem (the recommended harvest stage for Red Moon according to Mitchell Harty et al., 2008). Fruits were stored at 20 C over night a nd transported to the Food Science lab oratory the next morning. For the evaluation, two fruits of each type were selected and sliced into 15 g pieces (on average). Fruits were kept on ice until served to the panelist. In order to continuously serve fres h fruit throughout the day, two new fruits of each cultivar were sliced and served every 1.5 hours. Fruit from the four cultivars (s amples ) were each given a unique code and panelists were asked to taste each sample according to the code. Panelists rated the samples on a 9 point hedonic scale (1= dislike extremely, 2= dislike very much, 3= dislike moderately, 4= dislike slightly, 5 = neither like or dislike, 6= like slightly, 7= like

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91 moderately, 8= like very much, 9 = like extremely) for appearance ove rall acceptance, sweetness, flavor and firmness. Panelists were also asked to comment on each sample. Water and unsalted crackers were provided to cleanse the palate between each sample. A total of 74 panelists consiting of random visitors, staff and students, participated in the sensory evaluation. After the sensory panel was closed, aroma volatiles and SSC were collected on the remaining fruit that was used in the sensory evaluation, after the evaluation was completed. Fruit firmness was measured on re maining fruits. Statistical A nalysis A randomized complete block experimental design (RCBD) with four replications were used in fall 2006 and spring 2007, and three replications were used in fall 2007. Number of fruits per plot (n) ranged from three to 12 fruits. Data were analyzed using the GLM procedure (SAS Institute, Version 9, Cary, NC, U.S.A.). All data presented were subject ed to analysis of variance (ANOVA) and significant treatment means separated by Fishers least significant d ifference (P 0.05). Data was analyzed within season and harvest stage. Standard error (SE) values were calculated for each ethylene and respiration data point. Analysis for the melon sensory evaluation was done in a randomized complete block design (RCBD) with 74 pa nelists as the blocks. Means were separated using Tukeys HSD (P<0.05). Results Days to Harvest (DTH) Days to harvest (DTH) for Galia was generally later than the GT cultivars (Tables 4 2, 4 4, 4 6, 4 8). This indicated the speed in growth and deve l opment of the GT

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92 cultivars could be a beneficial trait for producers who require a fast growing or short season crop. DTH in fall 2006 and fall 2007 were 5 to 14 days less as compared to spring 2007 possibly due to greater temperatures compared to spring 2 007 (Table 4 10). Also, the shortest DTH occurred during fall 2007, where the greatest average, maximum and minimum temperatures as well as the greatest monthly solar radiation or photosynthetic photon flux (PPF) was recorded of all three seasons (Table 4 10). The increased temperature and light were due to the replacement of the greenhouse roof plastic, which was completed during summer 2007. Fruit Quality and Aroma Volatiles Stage ZG Stage ZG fruit at harvest indicated no differences in fruit quality for fall 2006 fruit (Table 4 2). Spring and fall 2007 fruit had some variation in weight and size, but most quality variables were similar among the cultivars (Table 4 2). All cultivars were generally firm at stage ZG, and in fall 2007, Elario also had the highest ethylene production (Table 4 2). Fruits harvested at stage ZG exhibited low volatile emissions at harvest for all cultivars, though increases in appeared after the five day storage period (at 20 C) in spring and fall 2007 (Table 4 3). Of th e 19 significant contributor (SC) compounds, there were only 11 to 15 found at harvest and 15 to 16 found after storage at stage ZG. The lowest number of SC compounds, with only Nos. 110, 15 (Table 4 1) was observed in fall 2006 at harvest In spring 200 7, there were 14 SC compounds (Nos. 1 4 and 6 15, Table 4 1) while fall 2007 had 15 SC compounds (Nos. 1 12, 1618; Table 4 1), although compound Nos. 16 and 17 were only present in significant amounts in Elario. Few

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93 differences were seen in the 38 iden tified volatiles among the cultivars at harvest (data shown in Appendix B -1, Tables 1 to 3, not needed for manuscript). After storage for five days in spring and fall 2007, TIV for stage ZG fruits increased over 200% for Galia and MG10183, while Elar io had no significant increase in TIV (Table 4 3). TIV for Galia at stage ZG were similar to harvest levels (Table 4 3). MG10183 was greatest in TIV after storage in spring 2007, but not in fall 2007, where Elario was greatest in TIV (Table 4 3). The increases in TIV after storage led to differences in 11 of the 38 identified volatile compounds between Galia and MG10183 in spring 2007, and differences in four compounds in fall 2007 (data shown in Appendix B -1, Tables 4 and 5, not needed for manuscript) Fruit firmness decreased from harvest in spring and fall 2007, to an average of 19 N and 13 N, respectively. Throughout the five days storage in spring and fall 2007 for stage ZG fruits, ethylene and respiration were generally low for all cultiv ars. In spring 2007, ethylene and respiration rates for Galia averaged 0.2 ng kg1 s1 and 7.1 g CO2 kg1 s1, respectively throughout storage. Also in spring 2007, ethylene and respiration rates for MG10183 averaged 0.3 ng kg1 s1 and 10.6 g CO2 kg1 s1, respectively, but also included a 69% increase in CO2 from day one to day two, followed by a decline. In fall 2007, ethylene and respiration rates were higher for both GT cultivars as compared with Galia (Fig. 4 1 A). Elario had an increase i n ethylene and CO2 on day two and then declined. Galia and MG10183 had a general increases in ethylene and CO2 throughout storage. The increases in both ethylene and respiration rates for both MG10183 and Elario at stage ZG, could represent the climacteric.

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94 Stage ZYG Fruit quality and aroma at stage ZYG was generally improved as compared with stage ZG as the fruits were in the process of ripening at this stage. Fruit quality factors, such as SSC increased while firmness decreased compared to stage ZG. Ethylene and respiration rates also increased from stage ZG as did aroma volatile emissions and additional differences were seen in the 38 volatiles among the cultivars. The total number of SC compounds at harvest ranged from 14 to 17. Fruit qualit y differences at stage ZYG within each season were minimal, variations occurred during fall 2006 and spring 2007 (Table 4 4). MG10183 was consistently firmer than Galia in fall 2006 and spring 2007. Differences in TIV were only observed in fall 2007, where Elario was greatest (Table 4 5). Elario was also over 500x greater in the compound methyl 2 -methyl butyrate, as compared to the other cultivars. SC compounds were lowest during fall 2006, with only 14, while spring 2007 and fall 2007 had 15 and 17, respectively. The SC compounds consistent in all cultivars were compound nos. 1 11, 1314 (Table 4 1), while No. 12 (ethyl isobutyrate) was not an SC in fall 2006 and No. 15 (cis 3 -hexenyl acetate) was not an SC in fall 2007. Fall 2007 also had compou nd No. 17, which was only significant in Galia and Elario and Nos. 18 and 19 were only significant in Elario. Among the 38 volatile compounds at harvest, there were only three differences among them in fall 2006 results. In spring and fall 2007, dif ferences among the 38 volatile compounds increased to 16 and 17 among the 38 compounds (data shown in Appendix B -2, Tables 1 to 3, not needed for manuscript). Following the five day storage period in spring and fall 2007, there were no differences in TIV among the cultivars (Table 4 5). There were 15 SC compounds ( Nos

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95 1 15, Table 4 1) present in all cultivars after storage in both seasons, however in fall 2007 ethyl 3 -(methylthio)propionate (No. 17, Table 4 1) was also an SC. During spring 2007, after s torage TIV increased for MG10183 while Galia was similar to its harvest TIV. Differences among the 38 compounds decreased to six between Galia and MG10183 (data shown in Appendix B -2, Tables 4 to 5, not needed for manuscript ). Fruit quality after storage in spring 2007 was similar to harvest results, except in firmness, which decreased and MG10183 remained a firmer fruit (20 N) after the five day storage period compared to Galia (4 N). After storage five days in fall 2007, TIV increased for a ll cultivars after storage as compared to harvest TIV. The compound methyl 2 -methyl butyrate increased and was similar among all cultivars. Differences among the 38 compounds remained similar to harvest levels with 15 compounds exhibiting variation (data s hown in Appendix B -2, Tables 4 to 5, not needed for manuscript) The only difference in fruit quality was again with firmness, as Elario (18 N) was firmer than MG10183 (10 N) and Galia (4 N). During the five day storage period in spring and fall 2 007, ethylene and respiration rates and patterns varied. In spring 2007, ethylene rates were averaged 0.6 and 0.7 ng kg1 s1 for Galia and MG10183, respectively throughout storage. Whereas during fall 2007, ethylene rates were o ver 200% greater and averaged 2 ng kg1 s1 for all cultivars throughout storage (Fig. 4 1 B). Respiration rates were similar in both seasons, averaging 10 to 12 g CO2 kg1 s1 for all cultivars during storage. As discussed earlier, the climacteric for the GT cultivars most l ikely began during stage ZG. As ethylene and respiration rates were increased at stage ZYG, the climacteric, most likely continued. The greatest ethylene and respiration rates were observed on either day one or two for all

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96 cultivars in both seasons, whic h was then followed by a continual decline through storage. Stages HS When results of stage HS fruit were compared to stage ZYG, it was found that most fruit quality factors and TIV remained the same except for firmness, which continue d to decrease and also for TIV during fall 2006, when MG10183 TIV increased from stage ZYG at harvest. The total number of SC compounds ranged from 14 to 18 at harvest and were similar to stage ZYG during all seasons, with the exception of fall 2007, where cis 3 hexenyl acetate was also an SC. Differences among the 38 compounds were less compared to stage ZYG, as there were differences in only five of the 38 compounds among the cultivars in fall 2006 and spring 2007, and only four in fall 2007 (data shown in Appendix B -3, Tables 1 to 3, not needed for manuscript) Ethylene rates at stage HS increased from stage ZYG in both spring and fall 2007 whereas respiration rates were similar to stage ZYG. Fruit quality at stage HS resulted in remarkable differences in SSC and firmn ess. MG1018 was consistently sweeter than Galia while Elario had the lowest SSC in fall 2007. The GT cultivars were also again firmer than Galia. TIV at stage HS were similar among all cultivars except for Elario, which was again greatest in T IV at harvest in fall 2007 (Table 4 7). Elario was also greatest again in methyl 2 -methyl butyrate at harvest. After three days storage in spring and fall 2007, TIV was similar among cultivars (Table 4 7). The SC compounds after storage in spring 2007 (stage HS) were similar to those at harvest. SC compounds in fall 2007 after storage were also similar to harvest SC compounds, except for compound No. 18 (isobutyl propionate), which was not a SC in

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97 Elario after storage. In spring 2007 there were no differences in the 38 volatile compounds between cultivars, however, in fall 2007 there were differences in 10 compounds after storage (data shown in Appendix B -3, Tables 4 to 5, not needed for manuscript ). Compared to harvest, TIV remained at similar l evels for spring 2007 fruits, while in fall 2007, Galia and MG10183 increased over 100% in TIV after storage (Table 4 7). After three days storage for stage HS fruits in spring and fall 2007 the average SSC of all cultivars was 10.1 Brix and 9.3 Bri x, respectively. Fruits firmness decreased from harvest for all cultivars and the GT cultivars grown in both seasons were firmer than Galia. Throughout the three day storage period during both seasons, ethylene rates were greatest on day one for Galia and MG10183 then declined during storage. In fall 2007, cultivar Elario was greatest in ethylene on day two (Figure 4 1). Respiration rates over the three day storage period were similar in both spring and fall 2007, data from fall 2007 is presented (Figure 4 1 C). The general respiration pattern during both seasons illustrated a decrease in CO2 by the end of the storage period. Stage FS By stage FS, fruit quality factors were similar to stage HS and TIV was greatest at stage FS for only fall 2006 and 2007 MG10183 fruits. St age FS TIV was similar to stage HS for all other cultivars. Fruit quality patterns within stage FS were also similar to stage HS as MG10183 was sweeter and firmer than Galia (Table 4 8). The number of SC compounds varied among seasons with 15 SC compounds in fall 2006 and spring 2007 and 16 in fall 2007. Differences in the 38 compounds were increased from stage HS and varied from 8, 6 and 10 through fall 2006, spring and fall 2007, respectively (data shown in Appendix B -4 Tables 1 to 3, not needed for manuscript). Interestingly of

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98 the 11 SC compounds in fall 2007, two compounds with a green aromatic scent (cis 6 nonen1 -ol and cis 3 -hexenyl acetate) were greatest in both GT cultivars. TIV at harvest were similar among the cultivars, although Elario still had the greatest amount of methyl 2 -methyl butyrate. After the two day storage period, Elario had the greatest TIV in fall 2007 and again, the greatest amount of methyl 2 -methyl butyrate (Table 4 9). SC compounds after storage were similar to harvest in both seasons, except for Elario, which increased 42% in TIV as compared to harvest TIV. There were differences in only two of the 38 compounds after storage in spring 2007 (data shown in Appendix B -4, Tables 4 to 5, not needed for manuscript) Each fruit quality variable ethylene and respiration rates prior to or after the two day storage treatment were similar among cultivars in spring 2007. After storage two days in fall 2007, there were differences in 11 compounds among the cultivars (data shown in Appendix B -4, Tables 4 to 5, not needed for manuscript), and cultivar Elario had the highest values for most of the compounds Also during fall 2007, differences after the two day storage period were observed i n fruit quality where MG10183 (11.3 Brix) was greater in SSC than Elario (8.7 Brix), but similar to Galia (9.6 Brix). Also, both Elario (15 N) and MG120183 (17 N) were firmer than Galia (9 N). There were no differences in ethylene or respi ration rates on day one or two of storage. All cultivars decreased in both ethylene and CO2 rates from day one (Figure 4 1 D). The decreases observed in ethylene and respiration could be indicative of these fruits in their post -climacteric stage.

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99 Sensory analysis A sensory evaluation was conducted on the Galia and GT muskmelons to further evaluate fruit quality and taste, and compare the panelist results to the fruit quality data. The sensory evaluation, which included Galia, both GT cultivars and Re d Moon (Figure 4 2) melon as a control, indicated that taste panelists preferred the appearance of Red Moon melon the most (Figure 4 3). Comments by panelists stated Red Moon flesh had great orange color. Overall acceptability (a general category of how much panelists liked the sample overall ) was greatest for MG10183. Both sweetness and flavor were greatest for MG10183 and Galia, which also were similar in SSC, which was above 11 Brix (Table 4 11). F irmness, though greatest in Newtons for Red Moon (Table 4 11), was less liked by panelists who rated MG10183 greatest followed by Red Moon. Galia was the least firm fruit and was also rated low, probably because of this. Although SSC was greatest for Galia, MG10183 and Red Moon (T able 4 11), panelists preferred MG10183 the most, followed by Gal ia. Comments by panelists indicated that Galias reduced firmness contributed to its low overall acceptance. However, the increased firmness of Red Moon also may have contributed to its low acceptance. Comments stated that Red Moon was too firm. Also, TIV was lowest for Red Moon (Table 4 11) and panelists commented that it had little flavor. This could indicate that the low TIV of Red Moon was not adequate enough to susta in a great acceptance, even though it had a high SSC. However a high TIV, as seen in Elario, which produced the greatest concentration of volatiles at every stage, except stage FS, did not result in good flavor as observed in the sensory panel. Elari o received the lowest score by panelists for flavor and sweetness, and correspondingly, also had the

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100 lowest SSC. Fallik et al. (2001) also reported that higher aroma does not necessarily mean higher consumer preference. In their study, the GT cultivar C8 had stronger aroma than cultivar 5080, but contained less sugar and was not preferred as much as 5080 by a taste panel. Th erefore the results from the spring 2008 sensory panel proved that high sugar, such as with Red Moon, does not necessarily equate to high consumer acceptance and that both increased aroma as observed in Elario and decreased aroma as observed in Red Moon also does not indicate high consumer acceptance. Firmness is another important issue as illustrated between Galia and MG10183 where both cultivars were sweet, but MG10183 was accepted over Galia due to Galia s texture, which was considered too soft. Although no storage test was conducted with Red Moon, Galia and the GT cultivars in spring 2008, previous rese arch demonstrated that Red Moon TIV decrease after storage (Mitchell Harty et al., 2008) while Galia and the GTs, as discussed in spring and fall 2007, generally increased in TIV following storage. This could also be an advantage to Galias consumer acceptance over Red Moon as the aroma of a food has an important influence in people choices (Lewinsohn et al., 2001). Differences occurred in 23 of the 38 identified compounds among the cultivars in the sensory analysis (Table 4 11). There were 13 SC c ompounds in all cultivars (Table 4 11). The compounds, isobutyl acetate, propyl acetate and ethyl 3 (methylthio)propionate were not SC compounds in spring 2008 in any cultivar. Also, isobutyl propionate was not an SC in Elario as in other seasons. The lower number of SC compounds during spring 2008 may be due to a slight variation in aroma volatile collection, due to the timing of the sensory analysis. In spring 2008, volatiles were collected on fresh, stage FS

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101 fruit that was used in the sensory evalua tion, after the evaluation was completed. This resulted in aroma volatiles being collected later in the day, after the fruit was chopped and remained out while the sensory evaluation was in session. Whereas in the previous seasons, aroma volatile collecti on was done immediately the day after harvest, with no extensive time elapsing once fruit was chopped. Therefore, these results stressed the importance of timely volatile data collection. Due to the volatile nature of these compounds, variation is often common in modifications to aroma volatile collection (Reguso and Pellmyr, 1998). However, measuring aroma volatiles on fresh fruit gives the closest representation as a consumer would find of what compounds are emitted. Of the SC compounds in spring 2008, benzaldehyde (almond scent), was greatest in Red Moon compared to the other cultivars. Red Moon also emitted high levels of, cis 6 nonen1 -ol (green, melon pumpkin scent) as did Elario. Isovaleronitrile (oniony, solvent scent) was once again a un ique SC to Galia. Throughout the stages and in the sensory panel, the aroma volatile analysis of the original Galia muskmelon and GT cultivars in this research revealed few distinctions among these cultivars, perhaps since they are derived from a lim ited germplasm base. TIV was mostly similar among cultivars Galia and MG10183 while Elario had greater in TIV at stages ZYG and HS at harvest, and stage FS after storage. The pattern of aroma development generally increased as fruits ripened for al l cultivars and continued to increase after storage (Fig. 4 3). Stage ZG had 12 SC compounds whereas this increased at stages ZYG, HS and FS where all had at least 14 SC compounds among all cultivars. The compound, isovaleronitrile was a unique SC to Galia. Isovaleronitrile is a nitrile with an oniony or

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102 solvent aroma (Khiari et al., 2003) that is thought to be derived from the amino acids leucine or isoleucine (Tieman et al., 2006). The compound isobutyl propionate was a unique SC to Elario. Isobutyl propionate is a methylpropanoate ester with a fruity or rum like odor (Smiley and Jackson, 2002). Furthermore, a SC unique to both GT cultivars was benzaldehyde, a compound with an almond aroma (Fischetti, 2000). Benzaldehyde, is also a SC compound found in red-fleshed melons such as Red Moon (Mitchell Harty et al., 2008). Of the 19 SC compounds found in the Galia and GT cultivars used in this study, all except isovaleronitrile of these have been reported in muskmelon. The SC compounds, isobut yl acetate, butyl acetate, 2 -methylbutyl acetate and hexyl acetate have been previously reported as the most abundant volatile compounds in other GT cultivars (Fallik et al., 2001). Additionally, Obando-Ulloa et al. (2008) reported that the GT cultivar Fa do was greatest in propyl acetate, methyl 2 -methylbutanoate as well as hexyl acetate, which is similar to what was measured in this study. Kourkoutas et al. (2006) found Galia contained greater levels of the acetate esters isobutyl, butyl, 2 -methylbutyl and hexyl acetate, than cantaloupe and honeydew. These compounds were also present in high concentrations in this study. Ethyl 3 (methylthio)propionate, a SC compound found in Galia and Elario only, is a sulfur compound with a fresh, melon like aro ma (Jordn et al., 2001), which may be important to the musky odor of muskmelon. Wyllie and Leach (1992) concluded that sulfur -containing compounds in the aroma volatiles of muskmelon were important and found that the Galia muskmelon used in their study had relatively intense amounts of another sulfur compound, 2 (methylthio)ethyl acetate. Ethyl 3 (methylthio)propionate, though present in this study, may not have been

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103 contributing much to the aroma. The SC compounds with high OVs, were benzyl acetate, ethyl 2 -methyl butyrate, ethyl isobutyrate, methyl 2 -methyl butyrate, ethyl butyrate, hexyl acetate, 2 -methylbutyl acetate and ethyl caproate (Table 4 11). The outcome of few differences in TIV and individual volatile compounds, especially between Galia and MG10183 may be attributed to the minimal difference between these cultivars or also, perhaps the sampling method may have to be altered. More samples as well as pooled samples may account for reduced variation and more significance, revealing a bet ter picture of the true differences. However, volatile collection using fresh samples, as in this study has probably resulted in a higher degree of true aromatic compounds identified. Not only are the amounts of individual volatile compounds important, but also the unique combinations of these volatiles that determine the aromatic properties (Thomson, 1987; Lewinsohn, et al., 2001) and add to the overall flavor. Perhaps the few differences observed in this research, such as isovaleronitrile as a SC compound in Galia only and the absence of ethyl 3 -(methylthio)propionate as a SC compound in MG10183 may be sufficient to account for a subtle difference in flavor and therefore, consumer preference. Based on this research, the compounds considered to be the most important to the high -quality original Galia muskmelon were benzyl acetate, ethyl 2 -methyl butyrat e, methyl 2 -methyl butyrate, ethyl isobutyrate 2 -methylbutyl acetate hexyl acetate, ethyl butyrate, ethyl caproate and cis 3 hexenyl acetate du e to their high OVs over a three season average. Additionally, isovaleronitrile and ethyl 3 ( methylthio)propionate may also be noteworthy; as isovaleronitrile was only a SC in Galia and ethyl 3 (methylthio)propionate was only a SC in Galia and Elario.

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104 A s research continues on aroma, collection methods are improved and more threshold levels are determined, other compounds will be revealed to play a greater role in the aromatic profile of Galia and other muskmelons, which will be useful in attainin g an exceptionally high-quality melon. Galia and GT muskmelons improved in overall fruit quality as the fruits ripened. Harvesting fruits prior to the climacteric phase, such as in stage ZG, resulted in increased firmness, but lower aroma. Harvesting at stage ZYG or later resulted in softer flesh, but there was increased aroma as fruits were at their climacteric peak. As illustrated in the sensory panel, sweetness, texture and aroma were key factors to the overall acceptance. Acceptable muskmelons must be sweet, but they must also be firm but not too firm (as with Red Moon). And good flavored melons must have sweetness, but also acceptable aroma, which must be at an acceptable level as was observed with fruits where aroma totals that were both high (Elario) and low (Red Moon) were not favored. Summary Galia muskmelon ( Cucumis melo L. var. reticulatus Ser.) is a world renowned cultivar, which has been popular for over 35 years and has generated its own market class of specialty melons called G alia type (GT) cultivars. The GT muskmelons are firmer that the original Galia but flavor has been compromised in breeding efforts to increase firmness. Since flavor is important to the quality of muskmelons, and includes the organoleptic traits of t aste, aroma and texture, it is important to determine factors that characterize a muskmelon with excellent quality, especially Galia, which is prized for its sweet, green, aromatic flesh. The high -quality Galia muskmelon and its GT relatives are there fore excellent candidates to study fruit flavor. To determine what has set Galia

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105 flavor apart from GT cultivars, this research focused on aroma, and sought to identify volatiles of the true Galia F1 hybrid. To evaluate aroma development, fruits were ha rvested at four stages: 1.) zero -slip, green (ZG); 2.) zero -slip, yellow -green (ZYG); 3.) half -slip (HS); and 4.) full -slip (FS). At each stage, quality factors (size, soluble solids content (SSC), firmness, pH, titratable acidity and aroma ), ethylene and respiration rates were measured. Galia muskmelon results were compared to the GT cultivar, MG10183 in fall 2006, spring and fall 2007; and to another GT, Elario in fall 2007. GC/MS and GC/FID verified 38 aroma compounds. Of these, 11 to 1 9 compounds significantly contributed to the aromatic profile, depending on stage and cultivar. Increases in aroma volatiles were observed as fruits ripened and after storage at 20C. Total identified volatiles ( TIV ) were lowest at stage ZG, where ethylene and respi ration rates were also lowest. Stage ZG TIV at harvest was similar for all cultivars during every season and fruit quality was also similar. As ethylene and respiration rates increased in stages ZGY, HS and FS, TIV also increased. The most differences in individual volatiles and TIV were seen during stage ZYG. Fruit quality differences were observed in firmness, where GTs were firmer than Galia. In spring 2008, A sensory evaluation was conduct ed on stage FS fruit at harvest and an additional cantaloupe cultivar, Red Moon which was marketed as the Perfect Melon, was included as a control and compared with Galia, MG10183 and Elario. Although Red Moon had the greatest firmness and had high SSC, taste panel preference was highest for MG10183, follow ed by Galia, then Red Moon. The least favorite cultivar was Elario. Based on this research, t he compounds considered to be the most important to high -quality Galia muskmelons were benzyl acetate, ethyl 2 -methyl butyrat e, methyl 2 -methyl bu tyrate, ethyl isobutyrate 2 -

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106 methylbutyl acetate hexyl acetate, ethyl butyrate ethyl caproate, cis 3 -hexenyl acetate isovaleronitrile, and ethyl 3 ( methylthio)propionate.

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107 Table 4 1. Volatile compounds considered to be significant contributors to t he aroma of Galia and GT cultivars, MG10183 and Elario fall 2006, spring 2007 and fall 2007. Ref No. Volatile compound Scent Odor threshold v alue (OTV) (ppb) Air/ water z OTV Ref. y 1 cis 6 nonen 1 ol waxy, melon, green, pumpkin 1 water 2, 6 2 ethyl caproate powerful fruity, pineapple, banana 1 water 6 3 benzyl acetate sweet, jasmine, apple, pear 0.04; 2270 air; ? 1,2 4 ethyl propionate sweet, fruity, ethereal 10 water 6 5 isobutyl acetate fruity 66 water 6 6 hexyl acetate fruity, green, pear (apple like) 2 water 6 7 ethyl butyrate fruity, pineapple, cognac 1 water 6 8 ethyl 2 methyl butyrate sharp, sweet, green, apple, fruity 0.1 0.3 water 6 9 2 methylbutyl acetate fruity 5 water 6 10 methyl 2 methyl butyrate sweet, fruity 0.25 water 6 11 amyl acetate bananas 7.5; 0.095 ?; air 5 12 ethyl isobutyrate sweet, rubber 0.1 water 6 13 propyl acetate nail polish remover 40 700 water 3 14 butyl acetate fruity 66 water 6 15 cis 3 hexenyl acetate powerful green, fruity, floral, banana melon 1.2; 7.8 water 10, 11 16 isovaleronitrile x oniony, solvent, fruity 3.2; 1000 water 8, 9 17 ethyl 3 (methylthio)propionate w fruity, pineapple, tropical 7 water 6 18 benzaldehydev bitter almond, almond 4.25; 350 3000 water 4, 9 19 isobutyl pr opionate u rum like, fruity 20 air 7 z, OTV as determined in air or water; ? = unknown.y, 1.) Waldhoff and Spilker. 2005; 2.) Burdock, 2005; 3.) SIS, 2007; 4.) Fischetti, 1994.; 5.) Ladd. Res. 2006; 6.) Leffingwell; 7.) Nagata, 1990 8.) Khiari et al., 2002, 9.) Buttery et al., 1991.x, Significant contributor in Galia only.w, Significant contributor in Galia and Elario only. v, Significant contributor in Elario and MG10183 only. u, Significant contributor in Elario only.

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108 Table 4 2. Stage ZG days to harvest and fruit quality means at harvest of Galia and Galia -type muskmelons z Non-significant (NS) or significant F test at P applicable, cultivar not grown. y, Mean separation by Fishers least significant difference test (P ) in fall 2007. Table 4 3. Stage ZG m eans of total identified volatiles (TIV) measured in ng gFW1 h1, from Galia and Galia -type muskmelons TIV at Harvest ( ng gFW 1 h 1 ) TIV after Storage ( ng gFW 1 h 1 ) Cultivar Fa06 z Sp07 z Fa07 y Sp07 Fa07 Galia 336 453 145 627 1 410 MG10183 553 450 352 2271 1187 Elario n/a n/a 903 n/a 1679 Significance z NS NS ** LSD 267 z Non -significant (NS) or significant F test at P grown. y, Mean separation by Fishers least significant difference test (P Cultivar Days to Harves t Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Fa06 z Sp07 z Fa07 y Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 38 56 28 1.53 1.15 0.91 154 142 128 143 126 118 35.9 29.4 22.9 MG10183 35 39 27 1.31 0.68 0.95 140 109 123 141 110 124 32.5 25.8 25.1 Elario n/a n/a 30 n/a n/a 1.29 n/a n/a 135 n/a n/a 141 n/a n/a 29.5 Significance z ** NS LSD0.27 NS LSD10.8 NS LSD14 NS NS LSD3.4 Cultivar Soluble s ol ids c ont ent (Brix) pH T otal titratable a cidity (T T A) Firmness (N) Ethylene (ng kg-1 s-1) Respiration (g CO2 kg-1 s-1) Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 10.5 9.5 6.3 7.17 6.55 6.15 0.11 0.14 0.14 39.8 46.2 42.0 0.21 0.05 0.2 9.18 4.63 11.7 MG10183 9.5 8.7 8.8 7.12 6.4 6.44 0.10 0.13 0.14 36.2 33.3 40.6 0.58 0.17 0.5 11.5 8.17 8.1 Elario n/a n/a 8.0 n/a n/a 6.21 n/a n/a 0.13 n/a n/a 51.0 n/a n/a 1.2 n/a n/a 10.7 Significance z NS NS NS NS NS N S NS NS NS LSD0.5 NS NS

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109 Table 4 4. Stage ZYG fruit quality means of Galia and Galia -type muskmelons Cultivar Days to Harvest Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Fa06 z Sp07 z Fa07 y Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 38 44 32 1.49 1.00 1.02 151 133 128 143 122 124 31.9 28 23.7 MG10183 35 38 29 1.24 0.65 1.20 135 111 135 136 107 132 30.5 25 28.5 Elario n/a n/a 29 n/a n/a 1.61 n/a n/a 150 n/a n/a 148 n/a n/a 34.8 Significance z 2.6 NS NS NS NS NS Cultivar Soluble solids c ontent (Brix) pH T otal titratable acidity (T T A) Firmness (N) Ethylene (ng kg-1 s-1) Respiration (g CO2 kg-1 s-1) Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 11.8 9.8 8.7 7.19 6.53 6.45 0.09 0.14 0.15 24.9 9.9 20.8 2.86 1.07 3.40 10.89 9.62 12.9 MG10183 12.5 10.6 8.9 7.35 6.77 6.35 0.10 0.12 0.14 32.1 36.1 35.1 5.31 1.87 3.80 16.61 13.3 11.9 Elario n/a n/a 6.8 n/a n/a 6.44 n/a n/a 0.12 n/a n/a 34.9 n/a n/a 5.70 n/a n/a 14.1 Significance z NS NS NS NS NS LSD0.02 ** ** NS NS ** z Non -significant (NS) or significant F test at P applicable, cultivar not gro wn. y, Mean separation by Fishers least significant difference test (P Table 4 5 Stage ZYG m eans of total identified volatiles (TIV) measured in ng gFW1 h1, from Galia and Galia -type muskmelons TIV at Harvest (ng gFW1 h1) TIV after Storage (ng gFW1 h1) Cultivar Fa06 z Sp07 z Fa07 y Sp07 Fa07 Galia 1646 2616 1387 2103 4877 MG10183 1322 1418 715 2246 4452 Elario n/a n/a 4790 n/a 4384 Significance z NS NS LSD 1870 NS z Non -significant (NS) F test at P 5 in fa ll 2006 and spring 2007; n/a= not applicable, cultivar not grown. y, Mean separation by Fishers least significant difference test (P

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110 Table 4 6 Stage HS f ruit quality means of Galia and Galia -type muskmelons Cultivar Days to Har vest Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Fa06 z Sp07 z Fa07 y Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 40 45 33 1.42 1.00 1.15 143 134 136 141 120 125 34.2 30 27.0 MG10183 34 40 31 1.62 0.70 1.38 150 114 143 151 108 139 36.4 25 29.6 Elario n/a n/a 31 n/a n/a 1.86 n/a n/a 154 n/a n/a 154 n/a n/a 35.3 Significance z ** ** LSD 1.2 NS NS LSD 0.6 NS LSD 21.3 NS NS Cultivar Soluble s olids c ontent (Brix) pH T otal titratable a cidity (T T A) Firmness (N) Ethylene (ng kg-1 s-1) Respiration (g CO2 kg-1 s-1) Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 12.3 9.8 9.4 7.2 6.73 6.46 0.10 0.13 0.12 21.5 10.9 11.7 2.84 2.48 3.6 12.2 12.3 10.5 MG10183 12.9 11.2 10.8 7.28 6.71 6.63 0.11 0.12 0.13 27.6 27.5 22.8 6.17 3.84 4.4 13.4 13.3 13.0 Elario n/a n/a 8.6 n/a n/a 6.67 n/a n/a 0.11 n/a n/a 23.8 n/a n/a 4.4 n/a n/a 13.4 Significance z NS ** LSD1.1 NS NS NS NS LSD0.02 N S ** LSD6.5 NS NS NS LSD1.5 z Non -significant (NS) or significant F test at P applicable, cultivar not grown. y, Mean separation by Fishers least significant difference test (P Table 4 7 Stage HS m eans, measured in ng gFW1 h1, of total identified volatiles (TIV), from Galia and Galia type muskmelons TIV at Harvest (ng gFW1 h1) TIV after Storage (ng gFW1 h1) Cultivar Fa06 z Sp07 z Fa0 7 y Sp07 Fa07 Galia 1624 1914 1545 2294 3756 MG10183 1656 2384 1007 2395 3208 Elario n/a n/a 5702 n/a 4439 Significance z NS NS LSD 3297 NS z Non -significant (NS) F test at P 5 in fall 2006 and spring 2007; n/a= not applicable, cultivar not grown. y, Mean separation by Fishers least significant difference test (P

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111 Table 4 8 Stage FS f ruit quality means of Galia and Galia type muskmelons Cultivar Days to Harvest Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Fa06 z Sp07 z Fa07 y Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 39 46 34 1.68 1.10 1.22 157 138 141 146 123 126 33.2 32 29.9 MG10183 38 43 32 1.44 0.80 1.10 147 119 131 145 112 129 32.9 28 27.9 Elario n/a n/a 32 n/a n/a 1.50 n/a n/a 144 n/a n/a 144 n/a n/a 32.9 Significance z NS ** LSD 0 .1 NS ** LSD 0.2 NS ** LSD 4.8 NS ** LSD 14.2 NS NS Cultivar Soluble solids c ontent (Brix) pH Total titratable a cidity (T T A) Firmness (N) Ethylene (ng kg-1 s-1) Respiration (g CO2 kg-1 s-1) Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 11.3 10.4 10 7.23 6.76 6.57 0.1 0.13 0.14 18.1 11.1 11.5 4.97 2.83 3.70 14.1 12.8 12.1 MG10183 13.1 12 11.2 7.26 6.76 6.61 0.09 0.11 0.13 24.1 22.7 20.4 5.76 3.34 5.10 13.6 12.7 10.8 Elario n/a n/a 9 n/a n/a 6.83 n/a n/a 0.09 n/a n/a 19 n/a n/a 3.90 n/a n/a 11.8 Significance z LSD1.3 NS NS NS NS LSD0.03 NS ** LSD6.8 NS NS NS NS z Non -significant (NS) or significant F test at P applicable, cultivar not grown. y, Mean separation by Fishers least significant difference test (P Table 4 9 Stage FS m eans of total i dentified volatiles (TIV) measured in ng gFW1 h1, from Galia and Galia -type muskmelons TIV at Harvest (ng gFW1 h1) TIV after Storage (ng gFW1 h1) Cultivar Fa06 z Sp07 z Fa07 y Sp07 Fa07 Galia 2190 1034 1616 2330 2645 MG10183 2446 1464 17 10 2828 2981 Elario n/a n/a 2700 n/a 3822 Significance z NS NS NS LSD 866 z Non -significant (NS) F test at P y, Mean separation by Fishers least significant difference test (P

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112 Table 4 10. Temperatures and photosynthetic photon fl ux ( PPF) during fall 2006, spring and fall 2007 of Galia and Galia type musk melons grown in a passivelyventilated greenhouse. Production month Average Temp. (C) Max. Temp. (C) Min. Temp. (C) Avg. Daily PPF (mol m2 s1) Max. Daily PPF (mol m2 s1) Aug. 2006 29.6 45.3 23.2 25.2 99.5 Sept. 2006 30.9 49.6 15.9 26.9 90.0 Oct. 2006 24.6 48.2 9.3 25.5 85.2 Feb/Mar. 2007 25.1 32.4 9.1 40.3 86.8 Apr. 2007 24.0 46.0 10.4 42.7 152 May. 2007 26.2 44.2 12.0 37.9 104 Jun. 2007 29.4 50.7 17.9 28.7 89.9 Aug. 2007 34.3 52.0 22.0 42.3 1937 Sept. 2007 29.6 54.0 20.4 308 1937 Oct. 2007 28.0 44.8 12.9 127 867

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113 Table 4 11. Stage FS m eans of soluble solids content (SSC, Brix), firmness (N) and aroma (ng gFW1 h1) of Galia, Galia -type and R ed Moon melons spring 2008. Fruit quality Galia MG10183 Elario Red Moon LSD (0.05) Z Soluble solids c ontent (Brix) 11.3 11.5 8.5 11.7 0.91 Firmness (N) 10.0 24.6 17.9 41.1 4.16 Aroma Compound Galia MG10183 Elario Red Moon LSD(0.05) Z propyl butyrate 2.04 2.42 2.74 0.63 tiglic aldehyde 0.31 1.51 2.14 0.58 0.67 4 methyl 1 cyclohexene 0.07 0.07 0.02 0.04 cis 3 hexen 1 ol 0.85 1.90 2.01 0.50 0.80 trans cyclodecene 0.08 0.16 0.50 0.50 0.10 cinnamyl acetate 0.13 0.34 0.42 0.24 0.19 2 methyl 1 butano l 14.7 3.72 3.98 1.01 2.82 furfuryl acetate 0.49 0.50 0.53 0.23 methyl isobutyrate 3.60 2.91 3.15 8.89 4.26 allyl methyl sulfide 0.67 0.08 0.47 0.39 butyl propionate 2.38 1.08 1.21 0.38 1.10 cyclooctene 0.70 0.68 1.07 1.13 isobutyl butyrate 3.32 2 .49 2.14 0.81 0.94 Benzaldehyde* GTs and R 1.90 4.46 4.56 7.15 2.41 3 phenylpropylacetate 2.30 9.71 5.40 1.60 4.27 methyl caprylate 11.5 4.07 6.59 6.35 methyl caproate 5.26 6.90 7.83 9.44 isobutyl propionate 8.09 5.08 7.89 4.89 2.42 ethyl 3 (methyl thio)propionate 1.76 2.29 2.84 0.19 0.74 heptyl acetate 8.80 27.6 25.8 1.62 7.23 phenethyl acetate 3.36 6.02 6.68 18.6 2.56 amyl acetate* G and GTs 19.6 36.6 32.7 6.3 .0 13.6 methyl butyrate 20.4 9.90 15.6 13.0 cis 6 nonen 1 ol* 24.4 98.5 117.0 172.5 5 7.6 Isovaleronitrile* G 14.1 0.07 0.08 0.08 5.27 ethyl caproate* 13.8 44.2 78.5 14.1 20.6 cis 3 hexenyl acetate 32.9 15.5 74.1 22.0 38.7 benzyl acetate* 0.70 1.2 32.4 59.6 ethyl propionate* 21.4 66.9 77.6 30.6 ethyl isobutyrate* 0.26 78.7 65.0 0.36 21.4 isobutyl acetate 32.4 10.8 21.9 9.33 9.66 propyl acetate 6.16 2.84 6.50 2.62 hexyl acetate* 24.2 18.0 27.0 7.76 ethyl butyrate* 39.1 8.00 12.4 28.7 17.3 butyl acetate* R 47.9 39.2 35.3 78.5 39.2 ethyl 2methyl butyrate* 12.9 55.7 67.1 15.6 2methylbutyl acetate* 138.1 64.0 96.9 33.2 methyl 2 methyl butyrate* 1636 1242 1581 11.7 407 Total Volatiles 1884 1877 2450 571 482 *Signif. Contributors 1726 1735 2224 472 489 z Mean separation by Fishers least significant difference test (P Units= ng gFW1 h1. *, denotes a significant contributor to the aroma G, significant contributor to Ga lia only. GT, significant contributor to Galia -types only. R, significant contributor to Red Moon only.

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114 Figure 4 1. Ethylene and respiration rates during storage at 20 C harvested at stages ZG, ZYG, HS and FS for Galia and GT muskmelons, fall 2007. A: ZG, Fall 2007 0 1 2 3 4 5 6 7 8 9 10 1 2 3 4 5 Days in Storage (20 C) Ethylene (ng kg-1 s-1) 0 2 4 6 8 10 12 14 16 18 g CO2 kg-1 s-1 B: ZYG, Fall 2007 0 1 2 3 4 5 6 7 8 9 10 1 2 3 4 5 Days in Storage (20 C) Ethylene (ng kg-1 s-1) 0 2 4 6 8 10 12 14 16 18 g CO2 kg-1 s-1 C: HS, Fall 2007 0 1 2 3 4 5 6 7 8 9 10 1 2 3 Days in Storage (20 C) Ethylene (ng kg-1 s-1) 0 2 4 6 8 10 12 14 16 18 g CO2 kg-1 s-1 D: FS, Fall 2007 0 1 2 3 4 5 6 7 8 9 10 1 2 Days in Storage (20 C) Ethylene (ng kg-1 s-1) 0 2 4 6 8 10 12 14 16 18 g CO2 kg-1 s-1 Galia MG10183 Elario G-Resp. M-Resp. E-Resp.

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115 Figure 4 2 Aroma volatile emissions of 17 SC compounds found in Galia, MG10183 and Elario, fall 2007, (Number (n) of fruits ranged from 1 to 12). 1= benzaldehyde, 2= i sovaleronitrile, 3= isobutyl propionate 4= ethyl 3 (methylthio)propionate 5= amyl acetate, 6= cis 6 nonen1 -ol, 7= ethyl caproate, 8= benzyl acetate, 9= ethyl propionate, 10= ethyl isobutyrate, 11= isobutyl acetate, 12 = propyl acetate, 13= hexyl acetate, 14= ethyl butyrate, 15= butyl acetate, 16= ethyl 2 -methyl butyrate, 17= 2 methylbutyl acetate A: Stage ZG, Fall 2007 0 50 100 150 200 250 300 350 400 450 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 Aroma volatile compounds ng gFW-1 h-1 B: Stage ZYG, Fall 2007 0 50 100 150 200 250 300 350 400 450 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 Aroma volatile compounds ng gFW-1 h-1 Galia MG10183 Elario C: Stage HS, Fall 2007 0 50 100 150 200 250 300 350 400 450 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 Aroma volatile compounds ng gFW-1 h-1 D: Stage FS, Fall 2007 0 50 100 150 200 250 300 350 400 450 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 Aroma volatile compounds ng gFW-1 h-1 Galia MG10183 Elario

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116 Figure 4 3. Galia, MG10183, Elario and Red Moon melons ( Cucu mis melo L.)used in the sensory panel, spring 2008. b b ab b b b a a a a b b c c b a bc a b bc 1 2 3 4 5 6 7 8 9 Appearance Acceptability Sweetness Flavor Firmness Hedonic Scale (1-9) Galia MG10183 Elario Red Moon Figure 4 4 Sensory evaluation results from fruit harvested at the recommended stage, FS for Galia, MG10183 and Elario; and the recommended harvest stage for Red Moon, which is when a crack begins at the abscission layer, spring 2008.

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117 CHAPTER 5 AROMA VOLATILE AND FRUIT QUALITY EVALUATION OF ANTISENSE ACC OXIDASE (CMACO 1) GALIA F1 HYBRID MUSKMELONS (Cucumis melo L. var reticulatus S er .) Galia muskmelon, a high quality specialty melon that fe atures a characteristic golden netted exterior and a sweet, green flesh, is renowned for its flavor (Aharoni et al., 1992; Karchi, 2000). In order to achieve the distinct flavor of Galia, it is best eaten when harvested at the fully ripe or full -slip st age (Karchi, 1979), thus its shelf -life is limited. Due to this limitation, Galia type (GT) cultivars have been bred, which are firmer than the original Galia muskmelon (Mitchell et al., 2007a). Although GT cultivars may have a longer shelf life tha n true Galia, many lack the high soluble solids content (SSC) and unique aroma of the original hybrid (Mitchell et al., 2007a; Cantliffe et al., 2001). Additionally, s everal methods have been used to extend the postharvest shelf life of Galia and GT muskmelons. They can be harvested early, at a green, pre -slip or half -slip stage when fruits are firmer, but this often results in poor sweetness and flavor (Fallik et al., 2001; Cantliffe and Shaw, 2002; Pratt. 1971). Muskmelons can also be stored at low temperatures (2.5 to 5 C) to maintain firmness (Asghary et al., 2005) or rinsed with hot water to reduce both fruit softening and decay development (Fallik et al., 2000; Lalaguna, 1998; Teitel et al, 1989). Galia muskmelons subjected to < 1.0 kGy of irradiation combined with a hot -water dip protected against decay and did not affect fruit quality (Lalaguna, 1998). Waxes have also been used to maintain internal and external melon fruit quality (Fallik et al., 2005; Aharoni et al., 1992). Sodium bic arbonate has been reported to reduce decay as well as maintain firmness (Aharoni et al., 1997). Moreover, a combination of hot water and a wax treatment with sodium bicarbonate 2007). Other postharvest treatments of GT muskmelons have included applications of hydrogen peroxide or

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118 thujaplicin, a chelating agent that inhibits microbial enzymes) treatments (Aharoni et al., 1994; Aharoni et al., 1993). Sto rage of Galia muskmelons in a controlled atmosphere of 10% CO2 and 10% O2 both with and without an ethylene absorbent decreased fruit softening and decay (Aharoni et al., 1993). The use of 1-methylcyclopropene (1 -MCP), an ethylene action inhibitor, supp ressed softening of Galia muskmelons at both green and yellow maturity stages (Ergun et al., 2006). Although these methods can be effective, many result in an extra step that producers and distributor s would most likely choose to avoid. T h erefore, instead of further complicating the postharvest handling process another means could be to modify the ripening pattern of the fruit itself. To do this, it is necessary to understand key features of the fruit such as the ethylene biosynthesis and its effect on fruit quality. The Galia mu skmelon is a climacteric fruit and has a burst of respiration concurrent with an autocatalytic production of ethylene (Abeles et al., 1992; Seymour and McGlasson, 1993). This is a defining feature of ripening i n fruits such as melons (Bower et al., 2002) and causes the fruit to ripen, abscise and soften very rapidly in most cases (Abeles et al., 1992). This has attributed to the short shelf life of Galia muskmelon Galia are best when harvested fully ripe, when the climacteric peak and abscission occur (Cantliffe and Shaw, 2002; Mitchell et al., 2007c) T herefore, knowledge about the ethylene biosynthetic pathway is important since it is essential to the fr ui t ripening process (Seymour and McGlasson, 1993) Ethylene biosynthesis follows the pathway from methionine via S adenosylmethionine (SAM) and 1 aminocyclopropane 1 carboxylic acid (ACC). The enzyme responsible for catalyzing the conversion of SAM to ACC, is ACC synthase (ACS) and the enzyme catalyzing ACC to ethylene is ACC oxidase (ACO) (Yang and Hoffman, 1984). However, the last step in

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119 the ethylene biosynthetic pathway can be inhibited using antisense technology and has been accomplished in tomato ( L ycopersicon e s culentum ) (Hamilton et al., 1990), Charentais and Vedrantais cantaloupes (Ayub et al., 1996; Guis et al, 1997 and 2000; Silva et al., 2004) and plums ( Prunus domestica L.) ( Callahan and Scorza, 2007). Furthermore, work by Nuez Palenius et al. (2006a) transformed the male parental line of Galia muskmelon (cv. Krymka) with an antisense ACC -oxidase gene (CMACO 1). Antisense Krymka fruits produced less ethylene and were firmer than wild -type (WT) fruits, yet soluble solids content (SSC) was similar to WT fruits (Nuez -Palenius et al ., 2006b). Nuez -Palenius et al. (2006a and 2006b) research provided an essential step towards the goal of improving the shelf life, while maintaining the high quality and flavor of the original Galia muskmelon. Nuez-Palenius (2006a and 2006b) determ ined that fruit quality factors such as SSC, TTA and pH were unaffected by the transgene and the ethylene inhibition by the antisense (CMACO 1) gene resulted in a firmer muskmelon. From that work, antisense ACC oxidase (CMACO 1) Galia (ASG) hybrid muskm elons were developed (Mitchell et al., 2007b). Although ASG muskmelons are firmer than Galia and have similar SSC, aroma volatile production associated with the ASG muskmelons has not yet been studied. This is important as changes in aroma also occur dur ing ripening and can affect fruit quality (Wang et al., 1996). The objective of this research was to determine whether or not adding an antisense ACC oxidase (CMACO 1) gene to the parental lines of Galia muskmelon give use to a longer shelf life Galia that retained all of the outstanding fruit quality characteristics. Materials and Methods Three experiments were conducted during fall 2006, spring and fall 2007. Seeds of the original Galia (Hazera Genetics, Israel) and antisense Galia (ASG) lines (ASxAS, ASxWT and WTxAS) were planted on 7 July 2006, 19 Jan. 2007 and 31 July 2007. ASG hybrid lines

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120 were produced from the Galia male parental line (cv. Krymka) that was previously transformed with an antisense ACC -oxidase gene (CMACO 1) by Nuez -P alenius et al. (2006a). Lines of antisense (AS) male parents were selfed and selected for the delayed ripening phenotype and a backcross 4 (AS BC4) female population was also produced. The AS T4 male and AS BC4 lines were crossed (ASxWT, WTxAS, ASxAS) and AS Galia F1 hybrid seed were produced in spring 2006 (Mitchell et al., 2007b). Seedlings were produced at the University of Florida, Gainesville, FL campus according to Mitchell Harty et al. (2008). Seedlings were grown in a Conviron plant growth chambe r (Controlled Env. Ltd.,Winnipeg, Manitoba, Canada) and fertilized once per week (after expansion of first true leaf) with Peters Professional All Purpose Plant Food (Spectrum Group, St. Louis, MO) at the rate according to Mitchell et al. (2007a). When se edlings had one true leaf, a polymerase chain reaction (PCR) analysis was used to identify the antisense seedlings with the transgene DNA was extracted from a 1.5 cm sample that was sliced from the youngest leaf tissue of each seedling The DNA extracti on method used was the Shorty buffer procedure for DNA isolation from the University of Florida, Hansen Lab, Nucleic Acid Isolation website (http://www.hos.ufl.edu/meteng/HansonWebpagecontents/NucleicAcidIsolation.html ) The PCR reaction was conducted in a DNA Thermal Cycler 480 (Applied Biosystems, Foster City, CA U.S.A. ) according to the parameters of Nuez -Palenius et al. ( 2006a) Also according to Nuez Palenius et al. (2006a ), electrophoresis of the amplified PCR products was done on a 1% agaros e gel and viewed by ultraviolet (UV) light. PCR analysis was completed on every putative transgenic Galia seedling. After transgenic seedlings were identified and all seedlings had three true leaves, they were transplanted on the 3 Aug. 2006, 27 Feb. and 15 Aug. 2007. All three trials were

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121 conducted in a saw tooth style, passively -ventilated greenhouse (TOP greenhouses, Ltd., Barkan, Israel), located at the University of Florida, Plant Science Research Education Unit located in Citra, FL. The plants wer e grown using commercial greenhouse muskmelon production techniques and nutrient requirements according to the recommendations of Shaw et al. (2001). Pollination was achieved via bumble bees from Class A research hives ( Bombus impatiens Natupol, Koppert Biological Systems, Inc., Romulus, MI). All flowers were tagged with the date of anthesis. Temperature and photosynthetic photon flux (PPF) at the canopy level were recorded daily at 30 -min. intervals by HOBO data loggers (Onset Comp. Corp., Bourne, MA). Within canopy temperatures were also recorded at 15 minute intervals by WatchDog data loggers (Spectrum Tech., Plainfield, IL). The monthly temperature averages were taken as an average of the within and at canopy readings. Insect pests and diseases were monitored weekly by scouting one plant from each plot per block. An integrated pest management (IPM) approach, which included the use of biological control and sprays, was used for management of arthropod pests in all three seasons and released at the rates according to Mitchell Harty et al. (2008). Fruit S election and Postharvest Treatments Fruits were harvested consistent with the methods Mitchell Harty et al. (2008) from 29 Sept. to 30 Oct. 2006; 10 May to 18 June 2007 and 2 Oct. to 26 Oct. 2007 at four different stages of ripening. Stage: 1.) zero-slip green (ZG): external skin still green in color with no abscission layer development; 2.) zero -slip, yellow -green (ZYG): external skin green and yellow, with no abscission layer development; 3.) ha lf -slip (HS): fruit abscising half -way; and 4.) full -slip (FS): fruit separates easily from the stem. During each harvest period, fruits were harvested each day, in the afternoon, and fruit weight and size (length and width) were recorded immediately.

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122 T he fruit was transported to campus and separated into two groups harvest and storage. In the harvest group, all postharvest variables were measured 12 hours after harvest. In the storage group, fruits were stored at 20 C and 85% relative humidity (RH). Storage days varied for the different stage fruits. This was done to be able to track the climacteric phase (ethylene and respiration rates were measured daily from each fruit while in storage) and collect fruit quality data at an appropriate edible time Stages ZG and ZYG were stored for five days, stage HS was stored for three days, and stage FS fruit was stored for two days. Only fruits harvested during spring and fall 2007 were subjected to a storage treatment. However, due to limited fruit number, line WTxAS was not included in the storage treatment. Ethylene and Respiration M easurements Ethylene and respiration measurements were measured on whole fruits from both H and S treatments consistent with methods and equipment as described by Mitchell Harty et al. (2008). After fruit were harvested, they were transported to Gainesville and stored at 20 C and 85% RH. The next morning, 12 hours after harvest, ethylene and respiration rates were measured from all fruits. Fruit Quality M easurements Fru it quality variables, which included firmness, SSC and flesh thickness were measured on fresh fruit according to Mitchell et al. (2007a) directly following storage for all treatments. Pulp firmness, SSC and flesh thickness data were recorded from a 2.5 cm slice, taken from the equatorial region of each fruit. These measurements were done using an Instron, refractometer and caliper. Pulp firmness was determined at two equidistant points on the equatorial region of each fruit slice using the Instron Univers al Testing Instrument (Model 4411-C8009, Canton, MA), which was fitted with a 50 kg load cell and an 11 mm convex probe with a crosshead speed of 50 mm min1. SSC (Brix) was measured with a temperature -compensating, handheld

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123 refractometer (Model 10430, Re ichert Scientific Instrument, Buffalo, NY) from fresh juice expressed from two pulp samples and a caliper (Digimatic Mycal, Mitutoyo, Japan) was used to measure flesh thickness from peel to cavity. The remaining pulp from the 2.5 cm slice was used for aro ma volatile collection and a quantity was frozen for Total Titratable Acidity ( T TA) an d pH measurements. Aroma volatile collection, TTA and pH measurements were all performed in accordance with methods and equipment as stated by Mitchell Harty et al. (200 8). In brief, methods for aroma volatile collections were performed according to Schmelze et al. (2003) and Tieman et al. (2006) with nonyl acetate as an internal standard. Aroma volatiles were collected from 100 g of fresh, chopped mesocarp pulp from eac h fruit that was inserted into Simex glass tubes (28 x 1.5 x 610 mm; Pegasus Glass). Air, via a vacuum pump, filtered through a hydrocarbon trap (Agilent Technologies, Palo Alto, CA), flowed through the tubes for one hour at 618 ml min1. Volatiles were collected on a Super Q column (30 mg Altech resin) and eluted with methylene chloride. Volatiles were separated on an Agilent Technologies DB 5 column (length x diam.: 30 x 0.25 mm) and analyzed on an Agilent Technologies 6890N (7683 Series) Gas Chromatograph (GC) (Agilent Tech., Inc., Santa Clara, CA). The GC was equipped with a flame ionization detector (FID). Retention times were compared with known standards and quantified with Agilent ChemStation software. Volatile peak identities were confirmed by an Agilent Technologies 5975 Gas Chromatograph/Mass Spectrometer (GC/MS). As a result of GC/MS, 38 volatile compounds were identified and standards were procured. Of the 38 compounds, 10 to 17 were considered to be significant contributors to the aroma depending on stage and season (Table 5 1). Significant contributors to aroma were identified as a result of dividing the concentration of the compound (determined with GC/FID)

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124 by its known odor threshold value (OTV), resulting in the odor value (OV) of the compound (Bauchot et al., 1998; Berger, 1995; Buttery 1993; Teranishi et al., 1991) Compounds with OVs greater than one were considered to be SCs to the aroma ( Bauchot et al., 1998). Odor threshold values (OTVs) were obtained through a literature se arch (Table 5 1). Statist ical A nalysis All trials were conducted in a randomized complete block design (RCBD) with three replications. Number of fruits per treatment (n) ranged from three to 12 fruits. Data were analyzed using the GLM procedure (SAS Institute, Version 9, Cary, NC, U.S.A.). All data presented were subject to Fishers least significant difference (P the two groups, harvest and storage, and analyzed by stage. A split -block experiment design with season as the main block and line as the split block were used for the combined analysis. An analysis of variance (ANOVA) was conducted using SAS (SAS Institute, Version 9, Cary, NC, U.S.A.). Results and Discussion Days to H arvest At stage ZG, only line ASxAS was harvested later than Galia, all other lines were harvested at similar times. By stages ZYG and HS, all ASG lines were harvested later than Galia (Tables 5 2, 5 3, 5 5). For stage FS fruit, there was a significant line x season interaction (Table 5 8). Although stage FS DTH varied among the seasons, generally, ASG lines remained on the vine longer than Galia, which is a significant feature of the delayed ripening characteristic. The delayed ripening and development of the abscission zone of the ASG fruit was also reported in antisense (AS) Krymka muskmelons (Nuez -Palenius et al., 2006b). The AS Krymka from Nuez Paleniuss research were used to develop these ASG lines (Harty et al., 2009). Throughout the three seasons, ASG lines remained on the vine an average of four days

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125 longer than Galia. However, this also depended on season a nd/or environmental conditions. Environmental influences commonly affect melon production and days to harvest (Bower et al., 2002). Generally, muskmelon growth was at an increased rate in the fall seasons, most likely due to higher temperatures at the onset of fruit set and development In fall 2007, fruits matured much earlier due to higher temperatures and solar radiation as compared with fall 2006 and spring 2007 (Table 5 8; Fig. 5 1). The increased temperature and light were due to the replacement of the greenhouse roof plastic, which was completed during summer 2007. Conversely, the growth rate of the spring 2007 crop was decreased due to cooler temperatu res at the onset of production and also to extended cloud cover during May due to wild fire smoke. During the spring, vast brush fires spread through Florida resulting in dramatic smoke levels near the greenhouse. Fruit Quality and Aroma Stage ZG Within the harvest treatment, there was no line x season interaction for any fruit quality factors or TIV at stage ZG over the three seasons (Tables 5 2 and 5 9). Fruit quality at stage ZG indicated few differences among lines, although Galia was a larger frui t. SSC was frequently lower for line WTxAS and fruit size also varied, but this could also be due to sampling variation. Fruits were firmest and SSC was lowest at stage ZG, which was analogous to fruit quality of other GT cultivars harvested at a green st age (Fallik et al., 2001). Ethylene and respiration rates were also lowest at this stage, demonstrating that stage ZG fruits were pre -climacteric as no significant increases in either gas occurred throughout the storage treatments. All lines had identica l significant contributor (SC) compounds, which generally consisted of at least 90% of the TIV, both at harvest and after storage at each stage. Aroma volatile production was lowest at stage ZG as compared with the other stages and the three season

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126 averag e resulted in only 11 SC compounds at harvest, stage ZG. The compounds that were not contributing significantly (amyl acetate, ethyl isobutyrate, propyl acetate, butyl acetate, ethyl 3 (methylthio)propionate and isovaleronitrile) were mostly esters with a fruity aroma. Other research has also reported lower amounts of aroma volatile compounds in immature cantaloupes as compared with mature fruits (Beaulieu, 2006; Beaulieu and Grimm, 2001 ; Wyllie et al., 1996b ). There were only two compounds with differences among th e 38 volatiles from all lines: methyl isobutyrate was greatest in Galia and cis 3 hexenyl acetate was greatest in line WTxAS. For stage ZG fruits that were stored for five days in spring and fall 2007 there were no differences in any postharvest variable s among the lines (data not shown). There was a seasonal difference for SSC only, where spring 2007 fruits were sweeter (8 Brix) than fall 2007 (6.2 Brix) fruits. There was a line x season interaction for firmness and ethylene production. For Galia and line ASxWT, fruits were firmer overall (15 N in spring 2007 and 8 N in fall 2007) and produced less ethylene (0.48 ng kg1 s1, spring 2007 and 1.75 ng kg1 s1, fall 2007 throughout storage) in spring 2007. Whereas line ASxAS fruits had similar firmness (9 N) and ethylene (0.5 ng kg1 s1) production in both seasons. Fruit firmness decreased after storage as compared with harvest levels in both seasons. The average respiration rate was 6.5 g CO2 kg1 s1 throughout storage for all lines, and no si gnificant increases in ethylene or respiration occurred throughout the storage treatments (data not shown). There was also a line x season interaction for TIV after storage (Table 5 10) After the five day storage treatment in fall 2007, Galia and line ASxWT had the greatest TIV and were significantly greater than their TIV at harvest (Table 5 10). Fallik et al. (2001) also observed an increase in aroma after storage of GT fruits harvested at a green stage. This did not happen for line ASxAS after st orage where TIV for line ASxAS was similar to its TIV at harvest. The total

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127 number of SC compounds increased as compared with those at harvest. There were 16 SC compounds for all lines in spring 2007, which included all SC compounds except isovaleronitr ile and these consisted of over 90% of the TIV after storage at stage ZG. In fall 2007, all lines had 15 SCs. Isovaleronitrile and cis 3 -hexenyl acetate were not considered SCs compound at this time. Stage ZYG Within the harvest treatment, fruit weight, size and SSC were similar among all lines. All ASG lines were firmer than Galia (Table 5 3). Compared to stage ZG, firmness decreased for all lines. The rate of fruit softening from stage ZG was greatest for Galia with a 53% decrease in firmness; whi le all ASG lines averaged a 31% overall decrease in firmness. Seasonal differences occurred for every variable, though fall 2006, generally had the largest and sweetest fruits; while fruits were firmest in both fall seasons. There was a line x season int eraction for TTA, pH, ethylene and respiration rates as well as TIV at stage ZYG (Tables 5 3, 5 4 and 5 9 ). TTA and pH values were inconsistent throughout the seasons, but ASG fruits generally had a lower TTA and higher pH than Galia. TIV increased for all lines from stage ZG, as did SSC (Tables 5 3, 5 4 and 59). Ethylene and respiration rates at stage ZYG, over the three seasons increased for all lines as compared with stage ZG (Table 5 4). However, ethylene rates for Galia at stage ZYG increased o ver 1500% from stage ZG double when compared to ASG lines, which had an average increase of 660%. The most differences in the volatile compounds among the lines were measured at stage ZYG, with differences in 20 of the 38 volatiles (Table 5 11) T his may be attributed to the increases in ethylene and respiration rates as the fruits ripened Other climacteric melons, such as the Charentais also exhibit an increase in aroma as they begin to ripen (Liu et al., 2004). However as Galia had greater amounts of the majority of the significant volatiles as compared with lines

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128 ASxAS and ASxWT, it appeared to be more aromatic. Bauchot et al. (1998 and 1999) reported that antisense ACO Charentais (Vedrantais) cantaloupes were less aromatic than the wil d type fruits with some esters reduced by 90% and total volatiles 60% to 85% lower. In this study, Galia had greater amounts of many of the volatiles at stage ZYG (Table 5 -11). Of the 17 SC compounds, there were 16 SCs in Galia and 15 in the ASG line s. The SC compound, ethyl 3 (methylthio)propionate was not significant in Galia nor in any ASG lines and isovaleronitrile was not a SC compound in any ASG lines at stage ZYG. Of the 16 SC compounds in the lines at stage ZYG, the five with the highest od or values (OVs), and therefore, contributing the most to the aroma profile, for Galia were benzyl acetate, ethyl 2 -methyl butyrate methyl 2 -methyl butyrate ethyl isobutyrate and, 2 -methylbutyl acetate (Table 5 12). The top five SC compounds in the A SG lines were also similar, though lines ASxWT and WTxAS had ethyl butyrate with the fifth highest OV (Table 5 12). Of the five volatiles with the highest OVs, Galia had 90% or more amounts of benzyl acetate, methyl 2 -methyl butyrate, ethyl isobutyrate and both Galia and line WTxAS were over 65% greater in 2 -methylbutyl acetate as compared with the ASG lines (Table 5 12). Due to the TIV line x season interaction, further analysis of each season revealed differences. In fall 2006, line WTxAS greatest ; in spring 2007, Galia was greatest and in fall 2007 Galia was only greater than line WTxAS (Table 5 9). Both lines ASxAS and ASxWT were only lower in TIV as compared with Galia in spring 2007; line ASxAS also lower than Galia in fall 2006. Duri ng fall 2006, at stage ZYG, there were differences in 26 of the 38 identified volatile compounds in fall 2006, stage ZYG fruits. Of the 26 volatiles with differences, line WTxAS produced the most in 19 volatiles ; both line WTxAS and Galia were great est in four, all lines except ASxAS were greatest in three. There were 14 SC compounds in

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129 all lines at stage ZYG, fall 2006 at harvest. The SC compounds, ethyl isobutyrate, ethyl 3 (methylthio)propionate and isovaleronitrile were not prominent at this stage. In spring 2007, TIV was great est for Galia (Table 5 9). Of the 38 volatile compounds, differences occurred in 16 compounds among the lines with Galia greatest in the majority. However, there were 16 SC compounds found in all lines at stage ZYG. These were similar to stage ZYG, fall 2006 with the addition of ethyl isobutyrate and isovaleronitrile. Ethylene and respiration rates were similar among all lines during this season. No differences in ethylene among Galia and the ASG lines in spring 2007 could be attributed to Florida wild fires, which spread through areas in close proximity to the greenhouse facilities. Smoke is well known to have ethylene as a ma jor component (Rodriguez, 1932). The ethylene produced from the wildfire smoke may have le d to early ripening of all fruits and thus no differences overall in ethylene production. Furthermore, after the fire/smoke event subsided, the plants were yellowed and damaged as a result. This demonstrated the sensitivity of the ASG muskmelons to stres s and emphasized that these ASG lines need to be produced under ideal conditions. Stressed plants can also produce more ethylene as a response (Srivastava, 2001; Morgan and Drew, 1997 ; McGlasson, 1970). This stress response was previously observed in stud ies on antisense ACC oxidase (CMACO 1) hybrids (TGH -AS 1 and TGH -AS 2) developed from a first generation transgenic male parent, which performed similarly to original Galia during an epidemic of severe powdery mildew ( Podosphaera xanthii (formerly Sphaer otheca fuliginea Schlech ex Fr. Poll.)) (Mitchell et al., 2007c). The TGH -AS 1 and TGH -AS 2 muskmelons had similar DTH to Galia in the diseased spring 2004 crop, while a fall 2004 crop was better managed for powdery mildew and differences were seen in D TH between Galia and the transgenic fruits remained on the vine an average of five days longer than Galia (Mitchell et al., 2007c).

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130 In fall 2007, there were no differences in TIV among Galia and lines ASxAS and ASxWT (Table 5 9). Of the 38 compounds, there were differences in 10 of which Galia was greatest in most (data not shown). There were 16 SCs for all lines; isovaleronitrile was not significant in any line at this stage. All ASG lines produced less ethylene and CO2 than Galia. Ethylen e rates at harvest increased from stage ZG for all lines except line ASxAS which remained at levels similar to stage ZG and further demonstrated its low ethylene production even as the fruit began to ripen. Respiration rates at harvest for all lines were similar to stage ZG (harvest) rates (Table 5 4) Stage ZYG fruits were stored for five days in spring and fall 2007. Differences were only seen in firmness among the lines where line ASxAS was firmer (6.3 N) after storage than lines ASxWT and Galia (bo th averaged 4 N). Compared to stage ZYG at harvest, SSC remained at levels similar to harvest while fruit firmness decreased for all lines. Seasonal differences occurred in fruit weight, size and firmness as fall 2007 fruits were 30% larger and firmer th an spring 2007 fruits. For all lines, t he overall average SSC was 8 Brix. Also at stage ZYG, after storage, there was a line x season interaction for ethylene, respiration and TIV For the ASG lines in fall 2007, ethylene and respiration rates were 70% and 25% lower, respectively, as compared with spring 2007 rates (Fig. 5 2). For Galia however, ethylene and respiration rates were 48% and 14% higher, respectively in fall as compared with spring 2007 (Fig. 5 2). In spring 2007, average ethylene evolut ion rates throughout storage were low and similar for all lines (Fig. 5 2). Respiration rates were lower for Galia and increases in CO2 were observed on day two for all lines (Fig. 5 2). Whereas in fall 2007, ethylene and respiration rates were greates t for Galia, especially at days one through three (Fig. 5 2). Increases in ethylene were observed on day two for Galia and ASxAS, while line ASxWT had

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131 an increase on day four of storage. Increases in respiration (CO2) were observed on day two for Ga lia and line ASxWT; and on day four for line ASxAS. These increases in CO2 could be representative of the climacteric occurring during stage ZYG. This is in contrast however, to antisense Charentais cantaloupes, which were reported to lack a climacteric rise in respiration (Bower et al., 2002). Although the ASG lines ASxWT and ASxAS in this study exhibited a slight rise in respiration during storage, respirations rates of these lines in fall 2007 were lower than Galia. The lack of consistency among Galia and the ASG lines in ethylene and respiration during both seasons after storage could be a result of the wildfire smoke during spring 2007. TIV after storage in spring 2007 TIV was similar among all lines, though increased after storage for the ASG lines, while Galia TIV remained at levels similar to harvest. Of the 38 volatiles, there were differences in only two among the lines hexyl acetate was greatest in Galia, while 2 -methyl 1 butanol was greatest in all ASG lines. All 17 SC compounds w ere present in all lines. In fall 2007, TIV was great est for Galia followed by line ASxAS. Line ASxWT was lowest in TIV (Table 5 10). Only the ASG lines increased in TIV compared to harvest TIV. There were differences in 15 of the 38 TIV compounds an d Galia was great est in all except two, where line ASxAS was also great est. There were 16 SC compounds for all lines, ethyl 3 (methylthio)propionate was not a SC at this time. Stage HS Analysis of the stage HS three season means at harvest indicated d ifferences in firmness, where lines ASxWT and ASxAS had the firmest fruits. The stage HS firmness decreased from stage ZYG for all ASG lines, while Galia remained the same. The continued firmness of these ASG lines at stage HS is an added highlight of t hese ASG hybrid muskmelons. Results from the antisense Krymka muskmelons (the original T0 antisense male parental line of the ASG lines)

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132 did not have firmer fruits at stage HS as compared with wild -type fruits (Nuez -Palenius et al., 2006b). Seasonal d ifferences occurred for all variables and there was a line x season interaction for TTA, pH and ethylene among the lines (Tables 5 5 and 5 6). TIV did not differ among the lines (Table 5 9), and there were differences in only six of the 38 volatile compou nds (data not shown). Compared to stage ZYG, TIV increased for all lines except Galia. Of the three season average, there were 16 SC compounds at stage HS in all lines. Ethyl 3 (methylthio)propionate was not a SC compound at stage HS. Further analys is of the line x season interactions indicated that ethylene rates varied over the seasons, though during fall 2006 and spring 2007, all lines had similar ethylene production rates. In fall 2007, l ines ASxAS and WTxAS produced less ethylene than Galia wh ile WTxAS produced the greatest amount of ethylene (Table 5 6). Compared with stage ZYG, stage HS ethylene production generally increased for all lines, except for Galia, in spring 2007, where there was a decrease in ethylene from stage ZYG. This decrease may also be related to no increase in TIV for Galia from stage ZYG to stage HS. The aroma produced as a result of the ripening process is associated with ethylene evolution and respiration rates (Ayub et al., 2008). Stage HS fruits were stored for three days in spring and fall 2007. Differences among the lines and seasons occurred for fruit weight and size, as line ASxWT had smaller fruits. There was a line x season interaction for firmness and TIV. In spring 2007, all lines had an average fruit firmness of 4 N, while in fall 2007, l ine ASxAS (10 N) had the firmest fruit as compared with Galia (6 N) and ASxWT (7 N). The firm ASG muskmelons in fall 2007 as compared with spring 2007 fruits at stage HS after storage not only continued to validate the problems with the spring crop, but also indicated additional potential for the ASG muskmelons at stage HS.

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133 As for TIV, in spring 2007, there were no differences in TIV among the lines (Table 510). There were differences in six of the 38 volatile com pounds among the lines in spring 2007, stage HS fruits after harvest (data not shown). All 17 SC compounds were present at stage HS after storage in all lines. In fall 2007 after storage, TIV were greatest for Galia and ASxWT (Table 5 10). There were d ifferences in 11 compounds among the lines (data not shown). There were 15 SC compounds after storage, both isovaleronitrile and ethyl 3 (methylthio)propionate were not among SC compounds in any line. Average ethylene production and respiration rates for stage HS fruit in both seasons throughout storage were 1.13 ng kg1 s1 and 10.9 g CO2 kg1 s1, respectively. Both ethylene and respiration rates generally decreased during storage (data not shown), indicating a decline in the climacteric during stage HS Stage FS At harvest, there were differences in fruit size, SSC, TTA and firmness among the lines and seasonal differences in most variables as well. There was also a line x season interaction for weight and length (Tables 5 7 and 58). Firmness decre ased from stage HS for all ASG lines and lines ASxAS and ASxWT were at a similar firmness to Galia. Line WTxAS was lowest in firmness and SSC; while ethylene and respiration rates were lowest for line ASxAS (Table 5 7). The lack of differences among th e ASG line and Galia at stage FS is again in contrast to full slip Krymka fruits, which were significantly firmer than the wild type (Nuez -Palenius et al., 2006b). TIV at stages HS and FS were similar for all lines. There were no differences in TIV among lines (Table 5 9). There were differences in only five of the 38 compounds at stage FS and there were 16 SC compounds at stage FS overall seasons (data not shown). E thyl 3 (methylthio)propionate was again, not a SC compound at stage FS. The SC comp ounds with the greatest OV were similar to those at stage ZYG for all lines (Table 5 12). Of these top five SC compounds, ethyl 2 -methyl butyrate and 2 -methylbutyl acetate were also considered important

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134 to the aroma of GT cultivars C8 and 5080 (Fallik et al., 2001). Additional volatiles reported in cultivars C8 and 5080 (Fallik et al., 2001), which were also among the 17 SC compounds in this study, were isobutyl acetate, butyl acetate, ethyl butyrate, ethyl hexanoate and hexyl acetate. Stage FS fr uits were stored for two days in spring and fall 2007. There were no differences in fruit quality variable among the lines after storage and only a seasonal difference for fruit width, where fall 2007 fruits (129 mm) were wider as compared with spring 2007 (115 mm) fruits. Fruit SSC averaged 9 Brix for all lines in both seasons after storage while firmness average 9N for all lines after storage in both seasons. There was a line x season interaction for TIV after storage. Compared to harvest TIV, after storage stage FS TIV increased for only Galia and line ASxWT. In spring 2007, TIV did not differ among lines; however fall 2007 TIV was greatest for line ASxWT (Table 510). In spring 2007 there were differences in seven of the 38 volatiles while in f all there were differences in four of the 38 compounds (data not shown). SC compounds totaled 17 after harvest for all spring 2007 fruit. There were only 15 SCs for ASG lines and 16 for Galia, fall 2007 fruit. Similar to fruit at harvest, ethyl 3 (meth ylthio)propionate was again not a SC compound in fall 2007 post -storage fruit, and isovaleronitrile was only an SC in fall 2007 GaliaBoth ethylene and respiration decreased in storage for all lines during both seasons (data not shown). The average ethyl ene production and respiration rates in both seasons were 2.57 ng kg1 s1 and 11.6 g CO2 kg1 s1, respectively among all lines in storage. For stages HS and FS ethylene and respiration rates at harvest were elevated in comparison to stages ZG and ZYG, though both decreased in storage, suggesting the end of the climacteric.

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135 Fruit harvest for all ASG lines was later than Galia at stages ZYG, HS and FS DTH for stage FS fruit over the three seasons provided the ASG lines a three to f ive day longer harv esting period. This indicated that the ASG lines ripened a t a slower rate than Galia, which is a common characteristic for antisense melons (Nuez -Palenius et al., 2006b; Ayub et al., 1996; Flores at al., 2001). Over the three seasons at stages ZG, HS and FS, fruit quality variables, ethylene and respiration rates and TIV were mostly similar among all lines both at harvest and after storage. Though some differences did occur such as at stage HS in fall 2007, where line ASxAS was the firmest line after storage. Individual volatile and TIV differences at stages ZG, HS and FS were few, as all lines produced ethylene and respiration at similar rates. The fruit quality and TIV similarities among the ASG lines and Galia at stage s ZG, HS and FS suggest tha t at these stages, ASG lines are identical in quality to the original Galia. The only difference was the length of time on the vine, which was longer for the ASG lines, particularly line ASxAS. The SC compounds at all stages, as well as those with the greatest OVs (Table 5 12), were mostly similar among all lines and overall seasons, except for ethyl 3 (methylthio)propionate and isovaleronitrile. The reduced amount of these volatiles could be an environmental influence as they were mostly absent in fa ll 2006 and spring 2007 at harvest. However, both of these volatiles usually increased after storage, most likely due to the continued ripening in storage. Ethyl 3 (methylthio)propionate and isovaleronitrile are considered important to the aroma of true Galia (Harty et al., 2009, unpublished). Also, TIV generally increased after storage at all stages, especially for Galia and line ASxWT. Both at harvest and after storage TIV remained the same for line ASxAS at stages HS and FS. Wyllie et al. (1996b ) also reported increases in total aroma volatiles after storage for Makdimon muskmelons harvested before and at full -slip.

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136 The majority of differences between the ASG and Galia were observed during stage ZYG. At stage ZYG, fruit quality in terms of S SC, T T A and pH was generally similar among Galia and lines ASxAS and ASxWT both at harvest and after storage Firmness, however, was greatest for ASG lines at harvest and after storage At harvest, e thylene rates were lowest for ASG lines at stage ZYG in fall 2006 and fall 2007, which could correspond to the firmer fruits. Low ethylene and firmer melon flesh was reported in antisense Charentais cantaloupes (Guis et al., 1997; Ayub et al., 1996) as well as antisense Kyrmka muskmelons (Nuez -Palenius e t al., 2006b). A s for aroma, TIV varied throughout the seasons at stage ZYG, though overall, Galia was greatest in TIV at stage ZYG at harvest, and after storage Galia had greater TIV in fall 2007. Low aroma volatiles have also been reported in antis ense ACO Vedrantais cantaloupes as compared with wild type cantaloupes (Bauchot et al., 1998 and 1999). W hile the reduction in aroma for the ASG lines at stage ZYG was not consistent in all seasons this could be due not only to environmental effects, but also due to high variation of the 38 volatile compounds (data not shown) This variation also resulted in no correlations between the volatiles and other fruit quality factors. In the future, additional samples and/or pooled samples may be necessary to obtain a more complete analysis of the individual lines to reduce variation and gain more information. Nonetheless, information obtained from these results suggested that fruits harvested at stage ZYG from line ASxAS exhibited the most potential for a l onger shelf -life high quality Galia muskmelon. Line ASxAS was consistently firmer and produced less ethylene than Galia at stage ZYG in fall 2006 and fall 2007 at harvest; and stage ZYG storage results demonstrated that line ASxAS remained a firmer fruit after five days at 20 C. TIV for line ASxAS was reduced compared to Galia at stage ZYG over all seasons, but this was not always

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137 found in every season Also, TIV for the ASG lines increased after storage indicating a more aromatic fruit. Altho ug h there was some reduction in aroma at stage ZYG for line ASxAS, overall, this line demonstrated positive results for an antisense ACC -oxidase Galia hybrid muskmelon with good fruit quality, flavor and a longer storage life. Summary Galia muskmelon ( Cucumis melo L. var. r eticulatus Ser.) is recognized for its aroma, an important quality and flavor component. Although Galia type (GT) cultivars are available that are firmer than Galia, they often have less flavor. In order to maintain the original Galia flavor while increasing firmness, three antisense ACC -oxidase (CMACO 1) Galia (ASG) hybrid lines were developed: ASxAS, ASxWT, WTxAS. The objective of this research was to evaluate fruit quality and aroma volatiles of Galia and ASG lines at diff erent stages of ripening and determine the effect of the genetic modification. During fall 2006, spring and fall 2007, fruits were harvested at four stages: stage 1.) zero-slip, green (ZG); 2.) zero -slip, yellow -green (ZYG); 3.) half -slip (HS); and 4.) ful l -slip (FS). Data were collected for fruit size, firmness, soluble solids content (SSC), pH and total titratable acidity (TTA). Aroma volatiles were collected from fresh pulp samples using the Super Q filter column method. GC/MS and GC/FID identified 38 aroma compounds, of which 18 were considered significant contributors to the aroma. Generally, aroma volatiles increased with maturity and after storage at 20C. At harvest, fruit firmness was greatest at stage ZG while SSC, ethylene and respiration rates and total identified volatiles (TIV) were low All lines were similar in TIV at stage ZG. At stage ZYG, ethylene and respiration increased and SSC was a minimum of 9 Brix for all lines while fruit firmness decreased. TIV was also lowest for lines ASxAS and ASxWT, while Galia had the great est TIV. The greatest differences were seen in the volatile compounds among the lines was at stage ZYG At stages HS and FS, ethylene evolution and respiration rates continued to be

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138 high, though individual volatile and TIV differences were few. After storage, there were no fruit quality differences at any stage among all lines in spring 2007. However, stage ZYG storage results from fall 2007 demonstrated that there is a potential for a longer shelf life ASG muskmelon as line ASxAS remained a firmer fruit after five days at 20 C. Fruit from the ASG lines remained on the vine an average of three to five days longer than Galia, suggesting a wider harvest window. Even though there were some differences in aroma volat iles at stage ZYG, it is recommended that line ASxAS be harvested at stage ZYG where SSC was acceptable and fruit firmness (for shipping) was greatest for fruits at harvest and after storage.

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139 Table 5 1. Odor detection threshold levels (OTV) of 17 the significant contributor aroma compounds of Galia and ASG muskmelons (adapted from Mitchell Harty et al., 2008). Volatile compound Scent Odor Threshold Value (OTV) (ppb) Air/ water z OTV Ref. y cis 6 nonen 1 ol waxy, melon 1 water 2, 5 ethyl caproate powerful fruity, pineapple, banana 1 water 5 benzyl acetate sweet, jasmine, apple, pear 0.04; 2 270 air; ? 1,2 ethyl propionate sweet, fruity, ethereal 10 water 5 isobutyl acetate fruity 66 water 5 hexyl acetate fruity, green, pear (apple like) 2 water 5 ethyl butyrate fruity, pineapple, cognac 1 water 5 ethyl 2 methyl butyrate sharp, sweet, green, apple, fruity 0.1 0.3 water 5 2 methylbutyl acetate fruity 5 water 5 methyl 2 methyl butyrate sweet, fruity 0.25 water 5 cis 3 hexenyl acetate po werful green, fruity, floral, banana, melon 1.2; 7.8 water 8, 9 amyl acetate bananas 7.5; 0.095 ?; air 4 ethyl 3 (methylthio)propionate fruity, tropical, grassy 7 water 5 ethyl isobutyrate sweet, rubber 0.1 water 5 propyl acetate nail polish remover 4 0 700 water 3 butyl acetate fruity 66 water 5 isovaleronitrile oniony, solvent, fruity 3.2; 1000 water 6, 7 z, OTV as determined in air or water; ? = unknown. y, 1.) Waldhoff and Spilker, 2005; 2.) Burdock, 2005; 3.) SIS, 2007; 4.) Ladd. Res. 2006; 5.) Leffingwell; 6.) Khiari et al., 2002; 7.) Buttery et al., 1991; 8.) Khiari et al.,1999; 9.) Belitz et al., 2004.

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140 Table 5 2. Stage ZG means of days to harvest and fruit quality variables for Galia and ASG lines at harvest fall 2006, spring and fall 2007. Line Days to h arvest Weight (kg) Length (mm) Width (mm) Flesh thickness (mm) Soluble solids c ontent (Brix) TTA pH Firmness (N) Ethylene (ng kg1 s1) Respiration (g CO2 kg1 s1) Galia 42 1.27 145 131 30.9 8.4 0.12 6.87 39.3 2.48 8.71 ASWT 40 0.93 130 118 24.8 7.8 0.11 6.85 41.0 0.16 7.35 WTAS 44 1.09 139 127 26.3 5.9 0.13 6.73 40.8 0.27 6.21 ASAS 45 1.23 135 126 27.8 8.0 0.11 7.01 41.9 0.17 6.04 LSD (0.05 ) z 2.9 0.24 9.7 7.4 2. 7 1.1 0.18 3.5 Season Fall 2006 39 1.40 146 137 31.7 9 .2 0.12 6.99 40.8 2.48 7.68 Spring 2007 57 1.13 139 125 28.0 7.8 0.12 6.70 39.0 0.13 4.40 Fall 2007 32 0.85 122 115 22.8 5.6 0.12 6.90 42.4 0.02 9.15 LSD (0.05) 6.0 0.25 6.3 6.5 2.7 1.5 0.3 0.4 4.2 z Mean separation by Fishers least significant di fference test (P y, Line x season interaction not significant for all variables.

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141 Table 5 3. S tage ZYG means of days to harvest and fruit quality variables for Galia and ASG lines at harvest fall 2006, spring and fall 2007. Line Days to h arvest We ight (kg) Length (mm) Width (mm) Flesh thickness (mm) Soluble solids c ontent (Brix) Firmness (N) Galia 38 1.16 138 129 27.5 10.1 17.9 ASWT 42 1.05 132 123 28.1 9.9 32.9 WTAS 42 1.18 136 129 30.4 9.2 22.2 ASAS 43 1.19 136 125 28.9 10.0 30.7 LSD (0.05 ) z 1.64 3.74 Season Fall 2006 41 1.37 147 137 30.8 11.4 30.9 Spring 2007 47 0.89 126 117 27.0 9.09 19.5 Fall 2007 38 1.17 132 128 28.2 8.91 27.6 LSD (0.05) 1.59 0.22 9.88 5.45 2.75 1.08 4.49 z Mean separation by Fishers least signific ant difference test (P

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142 Table 5 4. Stage ZYG means of significant line*season fruit quality parameters at harvest Line (L) TTA pH Ethylene (ng kg 1 s 1 ) Respiration (g CO 2 kg 1 s 1 ) Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 G alia 0.09 0.14 0.15 7.19 6.53 6.45 1.38 3.59 2.89 10.9 10 12.8 ASWT 0.09 0.11 0.09 7.37 6.99 7.37 1.75 1.75 1.39 11.8 10.7 8.67 WTAS 0.11 0.12 0.1 6.8 6.91 7.06 1.16 1.33 2.97 18.2 9.42 7.19 ASAS 0.11 0.11 0.11 7.17 6.86 7.17 0.77 1.2 0.86 10.1 10.1 7.1 9 LxS LSD (0.05) z 0.01 0.1 0.88 3. 1 z Mean separation for line x season interaction by Fishers least significant difference test (P

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143 Table 5 5 Stage HS means of days to harvest and fruit quality variables for Galia and ASG lines at harvest, fall 2006, spring and fall 2007. Line Days to Harvest Weight (kg) Length (mm) Width (mm ) Flesh thickness (mm) Soluble Solids Content (Brix) Firmness (N) Respiration (g CO2 kg1 s1 ) Galia 40 1.21 138 129 30.9 10.6 14.3 11.3 ASWT 42 1.30 142 130 29.5 9.9 21.8 11.3 WTAS 44 1.28 143 132 31.7 9.4 16.1 12.9 ASAS 43 1.27 140 131 30.0 10.2 18 .6 10.9 LSD (0.05)z 1.9 3.0 Season Fall 2006 42 1.61 154 144 34.7 11.3 21.8 13.6 Spring 2007 47 0.95 130 118 27.4 9.2 14.3 10.9 Fall 2007 37 1.23 139 129 29.5 9.5 17.0 10.2 LSD (0.05) 2.5 0.22 7.3 6.2 1.7 0.6 3.8 2.0 z Mean separatio n by Fishers least significant difference test (P

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144 Table 5 6. Stage HS means of significant line*season interaction of TA, pH and ethylene at harvest Line (L) TTA pH Ethylene (ng kg 1 s 1 ) Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Fa06 Sp07 Fa07 Galia 0.09 0.13 0.12 7.29 6.71 6.8 2.53 2.73 3.62 ASWT 0.1 0.12 0.11 7.17 6.72 7.2 3.67 1.81 4.56 WTAS 0.1 0.11 0.15 7.04 6.88 6.41 3.64 1.68 2.51 ASAS 0.11 0.11 0.11 7.19 6.96 7.18 2.47 1.9 1.33 LxS LSD (0.05) z 0.01 0.1 1.8 z Mean separation for line x season interaction by Fishers least significant difference test (P

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145 Table 5 7. S tage FS means of fruit quality variables for Galia and ASG lines at harvest fall 2006, spring and fall 2007. Line Width (mm) Flesh thickness (mm) Soluble solids c ontent (Brix) T T A pH Firmness (N) Ethylene (ng kg 1 s 1 ) Respiration (g CO 2 kg 1 s 1 ) Galia 133 31.9 10.6 0.13 6.91 14.0 3.78 13.1 ASWT 126 30.0 10.6 0.10 7.07 16.5 4.06 12.9 WTAS 125 27.3 8.2 0.11 7.10 11.0 4.06 13.2 ASAS 127 29.5 10.1 0.10 6.98 14.4 3.15 11.3 LSD (0.05) z 5.7 2.9 1.2 0.01 2.1 1.5 Season Fall 2006 139 33.0 11.5 0.10 7.16 17.0 4.51 13.8 Spring 2007 117 28.1 9.4 0.11 6.86 11.9 2.92 12.6 Fall 2007 128 27.9 8.7 0.11 7.04 13.1 3.89 11.4 LSD (0.05 ) 3.6 1.6 0.9 0.22 3.2 0.44 1.3 z Mean se paration by Fishers least significant difference test (P

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146 Table 5 8. Stage FS means of significant line*season interaction for days to harvest, weight and length Line (L) Days to harvest Weight (Kg) Length (mm) Fa06 Sp07 Fa07 Fa06 Sp07 F a07 Fa06 Sp07 Fa07 Galia 39 46 34 1.78 1.1 1.23 161 138 141 ASWT 44 47 37 1.29 0.83 1.3 142 126 143 WTAS 42 47 38 1.37 0.8 0.9 151 123 120 ASAS 46 49 38 1.35 0.87 1.23 148 126 138 LxS LSD (0.05) z 1.05 1.54 0.25 z Mean separation for line x sea son interaction by Fishers least significant difference test (P

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147 Table 5 9. Means of total identified volatiles (TIV) (ng g FW1 hr1) at harvest for Galia and ASG muskmelons harvested at stages ZG, ZYG, HS and FS in fall 2006, spring and fall 2007. Total Iden tified Volatiles (TIV) at Harvest (H) Line ZG z ZYG y HS FS Fa06 Sp07 Fa07 Galia 287 1646 2616 1387 1589 1636 ASWT 333 1586 581 1087 1775 2089 WTAS 310 2491 1087 776 2397 2015 ASAS 273 1201 526 973 1427 1855 LSD (0.05) x 425 425 425 Fall 2 006 290 2214 2760 Spring 2007 240 1391 1500 Fall 2007 372 1786 1437 LSD (0.05) 891 z ZG= stage zero -slip, green; ZYG= stage zero -slip, yellow/green; HS= stage half -slip; FS= stage full -slip. Stages ZG, HS and FS means are the three -s eason average. y There was a significant line x season interaction for stage ZYG only. LSD (0.05) for L x S interaction = 425 ng gFW1 h1. x Mean separation by Fishers least significant difference test ( P 0.05). Units= ng gFW1 h1. Table 5 1 0 Mea ns of total identified volatiles (TIV) (ng g FW1 hr1) after storage at 20 C for Galia and ASG muskmelons harvested at stages ZG, ZYG, HS and FS in spring and fall 2007. Total Identified Volatiles (TIV) after Storage (S) Line ZG z ZGY HS FS Sp07 Fa 07 Sp07 Fa07 Sp07 Fa07 Sp07 Fa07 Galia 627 1410 2103 4877 2294 3756 2330 2645 ASxAS 335 1722 1824 3034 1486 1389 2339 1742 ASxWT 467 680 2789 1436 1334 4324 1572 4240 WTxAS y 428 n/a 2249 n/a 2293 n/a 2086 n/a LSD (0.05) x 610 1107 1106 962 z ZG= stage zero -slip, green; ZYG= stage zero -slip, yellow/green; HS= stage half -slip; FS= stage full -slip. y WTxAS was only stored in Spring 2007. x Means separated using Fishers Least Significant Difference (LSD), P<0.05. Units= ng gFW1 h1.

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148 Table 5 1 1 Stage ZYG m eans, presented in ng gFW1 h1 of Galia and Antisense Galia (ASG) muskmelon aroma compounds measured at harvest fall 2006, spring 2007 and fall 2007. Aroma Compound Galia ASxAS ASxWT WTxAS Line z Season L*S propyl butyrate 0.98 0.75 0 .54 1.39 tiglic aldehyde 0.27 0.22 0.19 0.26 ** 4methyl 1cyclohexene 0.24 0.26 0.38 0.29 ** cis 3hexen 1ol 1.12 0.55 0.37 0.41 trans cyclodecene 0.98 0.48 0.20 0.25 cinnamyl acetate 0.32 0.33 0.29 0.15 2methyl 1butanol 0.37 0.16 0.17 0.19 furfuryl acetate 2.07 0.91 0.88 0.64 methyl isobutyrate 2.32 2.22 1.92 2.14 ** allyl methyl sulfide 1.51 0.39 0.35 0.72 ** ** butyl propionate 1.75 0.9 0.56 1.64 ** ** cyclooctene 0.94 1.42 0.63 1.45 isobutyl butyrate 1. 6 1.23 0.73 1.88 ** benzaldehyde 1.04 1.15 0.93 0.67 ** 3phenylpropylacetate 3.22 1.52 1.27 1.66 ** ** methyl caprylate 4.56 1.58 1.70 2.53 ** ** methyl caproate 12.8 9.49 7.24 9.42 ** isobutyl propionate 7.23 7.39 3.71 6.92 ** ethyl 3(methylthio)propionate 4.33 2.46 2.44 3.48 ** ** heptyl acetate 6.69 2.8 2.58 3.22 ** ** phenethyl acetate 12.2 6.01 3.72 1.8 ** ** ** amyl acetate^ 24.5 11.4 12.6 15.4 ** ** methyl butyrate 9.99 5.03 9.22 11.7 ** ** cis 6nonen1ol ^ 26.8 15.8 13.1 9.41 ** I sovaleronitrile ^ 4.68 1.56 1.31 1.97 ethyl caproate^ 48.6 24.2 22.5 26.7 ** ** cis 3hexenyl acetate ^ 19.3 6.14 10.1 16.1 ** ** ** benzyl acetate ^ 44.9 22.2 24.9 22.1 ** ** ** ethyl propionate ^ 40.9 33.1 42.7 28.7 ethyl is obutyrate ^ 33.9 15.5 12.5 9.67 ** ** ** isobutyl acetate ^ 289 181 322 337 ** propyl acetate ^ 127 67.5 120 86.1 ** ** hexyl acetate^ 57 27.6 66.2 114.7 ** ethyl butyrate ^ 56.6 44.6 42.4 42.8 ** ** butyl acetate^ 157 85.7 103 132 ethyl 2 met hyl butyrate ^ 41.4 37.9 34.6 38.2 2methylbutyl acetate ^ 328 167 243 357 ** ** ** methyl 2 methyl butyrate ^ 499 36.3 12.7 56.0 ** ** ** Total Identified Volatiles 1871 823 1123 1345 ** ** ** ^ Signif. Contributor (total) 1778 772 1074 1278 ** ** ** z, Significance among lines, seasons and line x season interaction. Mean separation by Fishers least significant d ifference test ( *= P 0.05, **=P 0.01).^, Denotes a significant contributor (SC) compound.

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149 0 10 20 30 40 50 60 Aug. 2006 Sept. 2006 Oct. 2006 Feb/Mar. 2007 Apr. 2007 May. 2007 Jun. 2007 Aug. 2007 Sept. 2007 Oct. 2007 Production Month and Year Temperature (C) 0 500 1000 1500 2000 2500 Solar Irradiation-PPF (mol m-2 s-1) Avg T Max T Min T Avg PPF Max PPF Figure 5 1. Average, maximum and m inimum temperatures and solar radiation (Photsynthetic Photon Flux (PPF)) for Galia and antisense Galia (ASG) produced in a passively ventilated g reenhouse, fall 2006, spring and fall 2007.

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150 CHAPTER 6 CONCLUSIONS The original Galia muskmelon ( Cucumis melo L. var. reticulatus Ser.) is recognized for its exceptional flavor and musky aroma. Newer Galia type (GT) muskmelons are firmer, but flavor has been compromised in breeding efforts to increase firmness. Th erefore a true Galia muskmelon with a longer shelf life is desir able to fully exploit flavor attributes while allowing longer distance shipping. The results of this research, which comprise d the development, selection and evaluation of an antisense ACC oxidase (CMACO 1) Galia F1 hybrid muskmelon, from fall 2006 through spring and fall 2007 indicated that developing a longer sh elf life true Galia muskmelon is possible Results from fal l 2006 indicated that ASG 1 muskmelons harvested at stages ZG, HS and FS exhibited many similarities to the original Galia muskmelon, especially in firmness and ethylene production. Only stage ZYG ASG 1 muskmelons from lines ASxAS and ASxWT demonstrated increased firmness and lower ethylene, however this only occurred when grown under optimal conditions ( such as good temperatures (Min: 18 C to Max: 35 C), minimal disease/insect pressure, and no wildfires or hurricanes ) There was also no difference in respiration among the ASG 1 lines and Galia at stage ZYG, or at any other stage. Other reports of antisense ACO Charentais cantaloupes report a lack of a climacteric rise in the antisense fruit compared with wild type fruit (Bower et al., 2002). The l ack of reduced respiration in the fall 2006 study suggested that a storage treatment was necessary to track gas emissions over time to better evaluate the effect of the reduced ethylene on respiration rates in the ASG 1 muskmelons. Nevertheless, fall 200 6 results of t he ASG 1 lines of ASxAS and ASxWT demonstrated the greatest potential for a longer shelf -life Galia muskmelon if harvested at stage ZYG and produced in an optimal environment. These ASG 1 lines exhibited similar fruit size (greater

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151 than 1 kg), similar SSC (ranged from 10.5 Brix to over 12 Brix) to the original Galia, yet were firmer than Galia at stage ZYG. The ASG 1 lines, ASxAS and ASxWT also produced less ethylene than Galia during fall 2006. Even though reduced ethylene was not observed again in spring 2007, most likely due to the environmental problems of wildfire smoke and reduced light intensity associated with that season, the ASG 1 lines ASxWT and ASxAS were again firmer than Galia in spring 2007 and had a later DTH than Galia. These results are similar to what Nuez -Palenius et al. (2006a and 2006b), Ayub et al. (1996) and Guis et al. (1997) reported in other antisense acc -oxidase melons in terms of firmer fruit, low ethylene and similar fru it quality (fruit size and SSC) to non antisense fruit types. Selections from the ASG material were made and used to continue research. Since flavor is important to the quality of muskmelons and includes the organoleptic traits of taste, aroma and texture (Goff and Klee, 2006), i t was important to determine factors that characterize a muskmelon which might possess excellent quality or poor quality. The high quality Galia muskmelon and its GT relatives are therefore excellent candidates to study fruit flavor. To determine why G alia flavor might be different from GT cultivars, this research also focused on aroma, in order to identify volatiles of the true Galia F1 hybrid. GC/MS and GC/FID verified 38 aroma compounds. Of these, 10 to 17 compounds significantly contributed to t he aromatic profile, depending on stage and cultivar. Increases in aroma volatiles were observed as fruits ripened and after storage at 20C. Based on this research, t he compounds considered to be the most important to high -quality Galia muskmelons were benzyl acetate, ethyl 2 methyl butyrat e, methyl 2 -methyl butyrate, ethyl isobutyrate 2 -methylbutyl acetate hexyl acetate, ethyl butyrate, ethyl caproate and cis 3 -hexenyl acetate due to their high OVs over a three -season average. OVs greater than one co ntribute the most to the aromatic profile and are

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152 obtained as a result of dividing the concentration of the compound (determined with GC/FID) by its known odor threshold value (OTV (Bauchot et al ., 1998; Teranishi et al., 1991). Additionally, isovaleronit rile and ethyl 3 ( methylthio)propionate may also be noteworthy; as isovaleronitrile was only a SC in Galia and ethyl 3 ( methylthio)propionate is a sulfur compound known to be impart to the musky aroma in muskmelons Wyllie and Leach (1992). The final objective of this research was to evaluate fruit quality and aroma volatiles of Galia and ASG lines at different stages of ripening and determine the effect of the genetic modification. During fall 2006, spring and fall 2007, fruits were harvested at four stages: stage 1.) zero -slip, green (ZG); 2.) zero -slip, yellow green (ZYG); 3.) half -slip (HS); and 4.) full -slip (FS). Results demonstrated that f ruit harvest for all ASG lines was later than Galia at stages ZYG, HS and FS D ays to harvest (DTH) for stage FS fruit over the three seasons provided the ASG lines a three to f ive day longer harvesting period. This indicated that the ASG lines ripened a t a slower rate than Galia, which is a common characteristic for antisense melons (NuezPalenius et al ., 2006b; Ayub et al., 1996; Flores at al., 2001). Over the three seasons at stages ZG, HS and FS, fruit quality variables, ethylene and respiration rates and TIV were mostly similar among all lines both at harvest and after storage. Generally, aroma vo latiles increased with maturity and after storage. Individual volatile and TIV differences at stages ZG, HS and FS were few, as all lines produced ethylene and respiration at similar rates. The fruit quality and TIV similarities among the ASG lines and G alia at stage s ZG, HS and FS suggest that at these stages, ASG lines are identical in quality to the original Galia. The only difference was the length of time on the vine, which was longer for the ASG lines, particularly line ASxAS.

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153 The greatest dif ferences between the ASG and Galia were observed during stage ZYG. At stage ZYG, fruit quality in terms of SSC, T T A and pH was generally similar among Galia and lines ASxAS and ASxWT both at harvest and after storage Firmness, however, was greatest for ASG lines at harvest and after storage At harvest, e thylene rates were lowest for ASG lines at stage ZYG in fall 2006 and fall 2007, which could correspond to the firmer fruits. Low ethylene and firmer melon flesh was reported in antisense Charentai s cantaloupes (Ayub et al., 1996) as well as antisense Kryrmka muskmelons (Nuez -Palenius et al., 2006b). A s for aroma, TIV varied throughout the seasons at stage ZYG, though overall, Galia was greatest in TIV at stage ZYG at harvest, and after storag e Galia had greater TIV in fall 2007. Low aroma volatiles have also been reported in antisense ACO Vedrantais cantaloupes as compared with wild -type cantaloupes (Bauchot et al., 1998 and 1999). After the five day storage period in spring and fall 20 07 for stage ZYG fruits, differences were only seen in firmness among the lines where line ASxAS had the firmest fruits (6.3 N) after storage than lines ASxWT and Galia (both averaged 4 N). TIV for line ASxAS was reduced compared to Galia at stage ZYG over all seasons, but low TIV for line ASxAS as compared with Galia was not consistent in every season. Also, TIV for line ASxAS muskmelons increased after storage, indicating a more aromatic fruit Although there was some reduction in aroma at stage ZYG for line ASxAS, overall, this line demonstrated positive results for an antisense ACC -oxidase Galia hybrid muskmelon with good fruit quality, flavor and a longer storage life. This research presents the problem concerning the short -shelf life of th e original Galia muskmelon and proposes the antisense ACC -oxidase (CMACO 1) ASG muskmelon as an improved alternative. However, there are both advantages and disadvantages to this alternative.

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154 The advantages are the delayed ripening characteristic, incr eased firmness at harvest and after storage as well as similar high quality to that of the original Galia hybrid. The delay in ripening would give growers an extended harvest period and allow ASG fruits to be harvested at the most advantageous stage, st age ZYG. Harvesting at stage ZYG resulted in firmer fruits and therefore, a longer shelf life. Finally, the quality of the ASG muskmelons is similar to the original Galia, a fruit marketed in the United States, Europe and the Mediterranean (Shaw et al. 2001; Rodriguez et al., 2001; Karchi, 2000. Although there are these several advantages of the ASG muskmelons, the disadvantages cannot be ignored. Disadvantages include the delayed ripening characteristic, environmental sensitivity, and the fact that this product is a genetically -modified organism (GMO). The delay in ripening, though also discussed to be advantageous, could also prove to be a burden on growers due to the longer production period. A longer growing season could result in additional expenses and time that growers may not have. The sensitivity of the ASG muskmelons, as observed in spring 2007 with the wildfire event and previously with pathogens such as powdery mildew, make production of ASG fruits a risky business. Lastly, GMO product s are often not favorably viewed by consumers due to their unknown health and environmental effects (Whitman, 2000). This resistance could result in a limited or no market for the ASG muskmelon. A non -GMO substitute for the ASG muskmelon could be newer Galia type (GT) cultivars such as MG10183 that was evaluated in this research. MG10183 demonstrated increased firmness as compared with the original Galia, yet was also preferred the most by consumers in a taste panel. In comparison with other GT cultivars, MG10183 was highly ranked in regard to fruit yield and quality (Mitchell et al., 2007a; Mitchell et al., 2006). Due to constant consumer complaints of fruits with little flavor, b reeders today are beginning to focus

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155 on quality in addition to yield for crops such as tomato (Causse et al., 2003). Melons, as well, are also being improved by breeders in terms of fruit quality a s well as shelf life and disease resistance ( Hoberg et al., 2003; Cohen et al., 2000). As breeder s employ traditional b reeding strategies that integrate improved quality factors the GM ASG muskmelon may merely serve as a scientific tool to investigate reduced ethylene in muskmelons.

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156 APPENDIX A CHAPTER 3 ANOVA TABLES Table A3 1. Source DF Mean square T1 T2 T3 T4 T rt. 2 345** 164* 675** 537** Rep 2 12.3 19.4 3.11 3.11 Error 4 13.8 23.1 1.61 4.11 and **, significant F test at P Table A3 2. Source DF Mean square T1 T2 T3 T4 Trt. 2 25.4* 10.3* 3.44 14.8 Rep 2 5.78 4.33 10.1 2 0.1 Error 4 1.94 0.67 1.78 13.4 and **, significant F test at P

PAGE 157

157 Table A 3 3 Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Trt. 4 15.8 136800 297 210 29.5 13.3** 39.3 10.4 18.7 Rep 3 15.1 34447 107 59.8 5.39 1.58 23.2 5.94 17.0 Error 12 4.93 81952 139 104 21.7 1.74 75.2 7.34 36.3 and **, significant F test at P .05 and P Table A3 4. Source DF Mean square Da ys to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Trt** 7 19.7** 81416 117.7 61.7 15.1 15.7** 108** 20.7** 89.1 Rep 3 3.6 111981 324.9 174 8.76 0.7 22.4 20.7 38.1 Error 21 5.64 72195 77.2 41.5 8.38 1.49 19.0 5.43 42.7 and **, significant F test at P Table A3 5. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Trt 8 56.9** 225892 308 147 37 5.64* 124** 87.9* 231 Rep 3 14.7 134477 153 124 15.6 2.09 2.4 43.7 58.2 Error 24 9.8 89809 78.5 68.7 20 1.69 28.5 20.7 216 and **, significant F test at P

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158 Table A3 6. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Trt 8 34.6* 191419 228 197 42.9 4.54* 35.2** 76.3 29.7 Rep 3 6.99 12873 4.48 34.2 4.01 2.13 27.4 11.5 3.37 Error 24 13.2 84512 140 86.4 20.6 1.46 10.2 12.5 19.7 and **, significant F test at P Table A3 7. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Line 3 37.0 0.14 289.7 198 53.7* 9.17* 58.4 0.11 12.3 Season 1 2346** 0.4* 373* 1213** 80.0* 20.5* 68.2 1.13* 368** L*S 3 11.7 0.06 96.2 68.6 10.2 2.38 22.3 0.09 22.4 Seas(Rep) 6 36.5 0.09 47.9 38.2 9.22 2.86 35.1 0.15 21.3 Error 18 16.1 0.08 140 82.9 13.1 2.27 57.1 0.16 30.9 and **, significant F test at P Tabl e A3 8. Source DF Mean square Days to harvest Weight Length Width Flesh SSC Firmness Ethylene Respiration Line 3 30.6** 0.05 123.3 82.1 3.88 2.49* 380** 25.0* 35.8 Season 1 259** 1.92* 3188** 3804** 142** 32.4* 1066** 19.8 185* L*S 3 1.78 0.03 72 41.7 3.72 4.84 11.8 16.6* 102** Seas(Rep) 6 3.61 0.03 98 31.3 5.32 1.44 10.9 3.46 27.1 Error 18 4.7 0.04 100.9 41.5 7.99 0.82 10.1 4.08 25.8 and **, significant F test at P

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159 Table A 3 9. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Line 3 18.4* 0.09 247** 60.9 16.4 5.48* 80.4** 5.35 25.7 Season 1 205** 3.07** 4052** 4910** 345** 42.6** 516** 95** 163* L *S 3 0.95 0.05 178* 40.7 18.5 3.45** 12.7 14.3* 55.1 Seas(Rep) 6 1.41 0.05 72.7 43.1 9.08 0.21 12.7 2.01 13.2 Error 18 4.13 0.04 44.2 38.1 16.9 1.04 14.2 3.44 29.1 and **, significant F test at P Table A3 10. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC Firmness Ethylene Respiration Line 3 35.4** 0.15 251** 170* 13.9 5.25** 24.3** 14.1 13.4 Season 1 145** 2.47 4010** 3885** 195** 29.5** 241** 132** 27.7 L*S* 3 6.92 0 .00 46.8 1.99 15.2 1.06 2.03 15.7* 14.9 Seas(Rep) 6 1.81 0.01 9.11 11.2 2.44 1.28 10.6 2.76 5.61 Error 18 2.03 0.03 42.8 36.9 8.7 0.77 3.77 7.87 6.31 and **, significant F test at P P

PAGE 160

160 Additional Tables for Chapter 3 Table A 1. Stage ZG external and internal color for Galia, Galia -type (MG10183) and grouped ASG lines, fall 2006. External Internal Line Lightness z Chroma y Hue angle x Lightness Chroma Hue an gle Galia 52.7 26.4 97.2 65.4 25.7 113.1 MG10183 55.5 36.3 102.1 76.4 24.9 112.2 ASWT (ASG 1) 47.4 22.4 93.1 74.6 23.5 111.7 WTAS (ASG 1) 50.6 38.3 107.0 71.8 25.8 112.2 ASAS (ASG 1) 55.1 23.5 102.3 73.2 25.2 111.3 ASAS2 (ASG 2) 46.8 16.9 101.9 71.3 28.9 112.5 LSD w NS NS NS NS NS NS z Lightness expressed on a scale from 0=black to 100=white. y Chroma expressed on a scale from 0=gray to 60. x Hue angles: red, 0; yellow, 90; green, 180; and blue, 270. w Means separated using Fishers Lea st Significant Difference; = significance at P<0.05; NS= not significant. Table A 2. Stage ZYG external and internal color for Galia, Galia type (MG10183) and grouped ASG lines, fall 2006. External Internal Line Lightnessz Chromay Hue anglex Ligh tness Chroma Hue angle Galia 52.6 32.18 97.8 73.2 25.1 110.6 MG10183 55.9 33.19 102.5 76.9 24.1 110.9 ASWT (ASG 1) 49.1 30.45 95.4 71.3 22.8 112.3 WTAS (ASG 1) 57.7 42.7 95.7 72.4 23.3 111.6 ASAS (ASG 1) 61.6 49.98 94.5 68.5 24.6 112.2 ASAS2 (ASG 2) 48.0 26.37 99.0 70.6 24.4 110.3 LSD w NS NS NS *2.95 NS 1.4 z Lightness expressed on a scale from 0=black to 100=white. y Chroma expressed on a scale from 0=gray to 60. x Hue angles: red, 0; yellow, 90; green, 180; and blue, 270. w Means sep arated using Fishers Least Significant Difference; = significance at P<0.05; NS= not significant.

PAGE 161

161 Table A 3. Stage HS external and internal color for Galia, Galia type (MG10183) and grouped ASG lines, fall 2006. External Internal Line Lightness z Chroma y Hue angle x Lightness Chroma Hue angle Galia 59.8 41.6 93.6 71.9 25.4 111 MG10183 66.9 45.9 92.9 75.1 22.4 112 ASWT (ASG 1) 63.6 42.9 89.1 71.1 22.1 112 WTAS (ASG 1) 63.4 40.2 91.8 73.3 24.1 112 ASAS (ASG 1) 63.8 45.1 92.4 70.0 25.9 112 ASAS 2 (ASG 2) 65.3 47.2 90.2 70.1 24.7 111 LSD w *3.64 NS NS *2.72 NS NS z Lightness expressed on a scale from 0=black to 100=white. y Chroma expressed on a scale from 0=gray to 60. x Hue angles: red, 0; yellow, 90; green, 180; and blue, 270. w Means separated using Fishers Least Significant Difference; = significance at P<0.05; NS= not significant. Table A 4. Stage FS external and internal color for Galia, Galia type (MG10183) and grouped ASG lines, fall 2006. External Internal Line L ightnessz Chromay Hue anglex Lightness Chroma Hue angle Galia 63.4 48.8 90.3 72.2 25.1 111 MG10183 56.9 37.9 94.7 73.4 23.4 112 ASWT (ASG 1) 62.2 41.6 91.7 68.6 23.0 112 WTAS (ASG 1) 60.1 45.4 85.9 70.2 25.8 111 ASAS (ASG 1) 60.7 38.3 96.1 68.8 24.5 1 12 ASAS2 (ASG 2) 65.4 48.9 90.5 71.1 23.3 111 LSD w NS NS NS NS NS NS z Lightness expressed on a scale from 0=black to 100=white. y Chroma expressed on a scale from 0=gray to 60. x Hue angles: red, 0; yellow, 90; green, 180; and blue, 270. w Means separated using Fishers Least Significant Difference; = significance at P<0.05; NS= not significant.

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162 APPENDIX B ADDITIONAL TABLES AND ANOVA TABLES FOR CHAPTER 4 Appendix B -1 Table B1 1: Stage ZG means of Galia and Galia type melon aroma co mpounds, fall 2006. Aroma Compound Signif.z Galia MG10183 propyl butyrate ns 0.88 0.61 tiglic aldehyde ns 0.17 0.13 4methyl 1cyclohexene ns 0.51 1.76 cis 3 hexen 1 ol ns 0.02 0.04 trans cyclodecene ns 0.19 0.05 cinnamyl acetate ns 0.06 0.03 2methyl 1butanol ns 0.01 0.00 furfuryl acetate ns 0.07 0.08 methyl isobutyrate ns 2.92 2.39 allyl methyl sulfide ns 0.04 0.34 butyl propionate ns 0.03 0.06 cyclooctene ns 0.10 0.30 isobutyl butyrate ns 0.05 0.09 benzaldehyde ns 0.31 0.53 3phenylpropylacetate ns 0.13 0.06 methyl caprylate ns 0.79 0.91 methyl caproate ns 5.32 4.31 isobutyl propionate ns 0.21 0.41 ethyl 3 (methylthio)propionate ns 0.66 1.38 heptyl acetate ns 0.77 1.14 phenethyl acetate ns 0.28 1.08 amyl acetate ns 1.72 2.56 m ethyl butyrate ns 7.52 11.1 cis 6nonen1ol ^ ns 6.81 11.9 isovaleronitrile ns 0.10 0.04 ethyl caproate ^ ns 10.2 7.27 cis 3 hexenyl acetate ns 2.08 9.83 benzyl acetate ^ ns 2.73 6.16 ethyl propionate ^ ns 33.5 21.7 ethyl isobutyrate ns 0.01 0.01 isobutyl acetate^ ns 103 162 propyl acetate ns 38.8 59.2 hexyl acetate ^ ns 10.5 24.2 ethyl butyrate ^ ns 21.9 25.2 butyl acetate ns 12.3 34.2 ethyl 2 methyl butyrate ^ ns 21.9 12.8 2methylbutyl acetate ^ ns 48.0 115 methyl 2 methyl butyrate ^ ns 25.0 24.6 Total Volatiles ns 360 553 ^ Sig. Contributors ns 283 420 z, ns, non significant F test at P

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163 Table B1 2: Stage ZG m eans of Galia and Galia type melon aroma compounds, spring 2007. Aroma Compound Signif. z Galia MG10183 propyl butyrate ns 0.47 0.54 tiglic aldehyde ns 0.01 0.00 4 methyl 1 cycloh exene ns 0.12 0.03 cis 3 hexen 1 ol ns 0.30 0.18 trans cyclodecene ns 0.16 0.09 cinnamyl acetate ns 0.32 0.12 2 methyl 1 butanol ns 0.75 0.03 furfuryl acetate ns 0.51 0.47 methyl isobutyrate ns 0.08 0.11 allyl methyl sulfide ns 0.09 0.38 butyl prop ionate ns 0.22 0.24 cyclooctene ns 0.20 0.26 isobutyl butyrate ns 0.23 0.25 benzaldehyde ns 1.50 0.21 3 phenylpropylacetate ns 0.75 0.42 methyl caprylate ns 0.87 1.55 methyl caproate ns 2.73 4.30 isobutyl propionate ns 1.18 1.89 ethyl 3 (methylthio )propionate ns 1.51 2.31 heptyl acetate ns 1.63 2.05 phenethyl acetate ns 0.10 0.09 amyl acetate ^ ns 6.52 8.53 methyl butyrate ns 8.18 0.19 cis 6 nonen 1 ol ^ ns 1.82 2.23 isovaleronitrile ns 0. 03 0.01 ethyl caproate ^ ns 15.6 8.01 cis 3 hexenyl acet ate ^ ns 16.4 19.4 benzyl acetate ^ ns 19.6 20.3 ethyl propionate ^ ns 36.4 40.4 ethyl isobutyrate ^ ns 38.9 8.05 isobutyl acetate ns 34.3 19.5 propyl acetate ^ ns 63.1 74.8 hexyl acetate ^ ns 31.8 44.5 ethyl butyrate ^ ns 20.3 27.2 butyl acetate ^ ns 27.1 91.6 ethyl 2 methyl butyrate ^ ns 27.6 15.2 2 methylbutyl acetate ^ ns 89.0 51.2 methyl 2 methyl butyrate ^ ns 2.47 3.86 Total Volatiles ns 453 450 ^ Sig. Contributors ns 426 408 z, ns, non significant F test at P

PAGE 164

164 Table B1 3: Stage ZG m eans of Galia and Galia type melon aroma compounds, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 0.36 0.16 0.89 tiglic aldehyde 0.27 0.04 7.79 4 methyl 1 cyclohexene 0.04 0.44 0.12 cis 3 hexen 1 ol 0.29 0.31 0.61 trans cyclodecene 0.26 0.12 0.26 cinnamyl acetate 0.20 0.17 0.26 2methyl 1butanol 0.21 0.03 0.61 furfuryl acetate 0.17 0.18 0.36 methyl isobutyrate 0.39 0.07 0.07 allyl methyl sulfide 1.31 0.22 0.11 butyl propionate 0.33 0.17 0.44 cyclooct ene 0.03 0.20 0.29 isobutyl butyrate 0.32 0.16 0.58 benzaldehyde 1.30 2.38 8.64 3phenylpropylacetate 0.36 0.19 0.64 methyl caprylate 0.48 1.27 2.29 methyl caproate 1.57 3.07 9.06 isobutyl propionate 0.59 0.54 3.11 ethyl 3 (methylthio)propion ate 5.97 1.83 13.9 heptyl acetate 0.78 0.88 1.75 phenethyl acetate 0.75 2.86 8.63 amyl acetate 3.09 3.10 3.26 methyl butyrate 0.11 0.34 19.4 cis 6nonen1ol 5.30 0.39 4.06 14.8 isovaleronitrile 0.17 0.09 0.23 ethyl caproate 3.98 6.25 35.9 c is 3 hexenyl acetate 0.40 0.29 1.58 benzyl acetate 7.95 36.0 89.2 ethyl propionate 2.34 2.62 39.1 ethyl isobutyrate 0.24 3.35 24.2 isobutyl acetate 20.7 53.3 71.3 propyl acetate 11.0 22.0 26.4 hexyl acetate 17.2 21.1 45.6 ethyl butyrate 18.4 28.5 123 butyl acetate 7.11 30.3 39.8 ethyl 2 methyl butyrate 3.90 10.8 41.2 2methylbutyl acetate 29.8 81.5 266 methyl 2 methyl butyrate 2.29 12.8 16.5 Total Volatiles 145 352 903 Signif. Volatiles 135 338 835 z Mean separation by Fishers least significant difference test (P

PAGE 165

165 Table B1 4: Stage ZG means of Galia and Galia type melon aroma compounds following storage for 5 days at 20C, spring 2007. Aroma Compound Signif. z Galia MG10183 propyl butyrate ns 1.24 1.35 ti glic aldehyde ns 0.02 0.02 4 methyl 1 cyclohexene ns 0.04 0.03 cis 3 hexen 1 ol ns 0.26 0.75 trans cyclodecene ns 0.22 0.38 cinnamyl acetate ns 0.31 0.32 2methyl 1butanol ns 0.12 0.32 furfuryl acetate 0.79 1.58 methyl isobutyrate ns 0.08 0.06 a llyl methyl sulfide ns 0.12 1.83 butyl propionate ns 0.83 0.56 cyclooctene ns 0.30 0.46 isobutyl butyrate ns 0.62 0.54 benzaldehyde ns 0.34 0.14 3phenylpropylacetate ns 0.93 1.43 methyl caprylate ns 2.54 10.2 methyl caproate ns 8.03 4.95 isobutyl propionate 0.92 2.57 ethyl 3 (methylthio)propionate 9.06 3.94 heptyl acetate ns 1.78 3.55 phenethyl acetate ns 0.30 0.58 amyl acetate* 6.0 13.4 methyl butyrate ns 10.7 2.39 cis 6 nonen 1 ol ns 2.11 3.75 isovaleronitrile ns 0.04 0.05 ethyl cap roate* ns 21.3 5.15 cis 3hexenyl acetate* 19.5 180 benzyl acetate* 12.7 77.1 ethyl propionate* ns 49.7 68.4 ethyl isobutyrate* ns 15.5 8.52 isobutyl acetate* 14.8 68.8 propyl acetate* 60.0 228 hexyl acetate* ns 41.1 84.7 ethyl butyrate* ns 89.4 70.4 butyl acetate* ns 53.9 166 ethyl 2methyl butyrate* 47.5 20.6 2methylbutyl acetate* ns 125 370 methyl 2 methyl butyrate** 22.3 855 Total Identified volatiles 627 2271 Signif. contributors 568 2053 ns, and **, not significant and significant F test at P 0.05 and P

PAGE 166

166 Table B1 5: Stage ZG means of Galia and Galia type melon aroma compounds measured after 5 days storage at 20C and harvested at stage ZG, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 2 .31 5.92 2.63 4.74 tiglic aldehyde 0.01 0.02 0.03 4methyl 1cyclohexene 0.21 0.12 0.27 cis 3 hexen 1 ol 0.59 0.23 0.59 trans cyclodecene 0.66 0.31 0.45 cinnamyl acetate 0.27 0.23 0.32 2 methyl 1 butanol 0.58 0.29 0.52 furfuryl acetate 1.61 1 .64 2.30 methyl isobutyrate 0.01 0.02 0.09 allyl methyl sulfide 2.00 2.64 3.25 butyl propionate 3.22 1.55 2.20 cyclooctene 1.18 0.35 0.89 isobutyl butyrate 3.70 1.19 2.82 benzaldehyde 4.61 3.15 5.22 11.5 3 phenylpropylacetate 2.44 1.52 4.98 m ethyl caprylate 2.02 1.74 3.23 methyl caproate 5.01 2.67 5.69 isobutyl propionate 8.33 5.78 8.30 ethyl 3 (methylthio)propionate 7.91 5.78 6.67 heptyl acetate 2.81 1.83 3.71 phenethyl acetate 17.6 31.6 20.4 amyl acetate 7.41 8.06 10.8 methyl butyrate 3.18 10.7 0.71 0.59 pentyl acetate 21.3 14.3 22.9 cis 6 nonen 1 ol 5.81 4.84 50.5 isovaleronitrile 0.12 0.17 0.11 ethyl caproate 23.1 8.63 12.4 cis 3 hexenyl acetate 0.85 0.83 2.98 benzyl acetate 98.2 39.3 96.1 201 ethyl propionate 55 .2 17.1 96.4 ethyl isobutyrate 16.2 0.45 23.8 isobutyl acetate 41.9 31.8 75.3 propyl acetate 209 231 161 hexyl acetate 170 100 246 ethyl butyrate 203 134 187 butyl acetate 179 228 107 ethyl 2methyl butyrate 87.0 39.2 94.2 2 methylbutyl ace tate 242 191 252 methyl 2 methyl butyrate 30.1 13.2 54.1 Total Volatiles 610 1410 1187 1679 z Mean separation by Fishers least significant difference test (P 0.05).

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167 Appendix B -2 Table B2 1: Stage ZYG means of Galia and Galia type melon aroma compounds, fall 2006. Aroma Compound Signif. z Galia MG10183 propyl butyrate ns 0.27 0.24 tiglic aldehyde ns 0.55 0.48 4 methyl 1 cyclohexene 0.51 1.75 cis 3hexen 1ol ns 0.32 0.11 trans cyclodecene ns 0.14 0.15 cinnamyl acetate ns 0.11 0.03 2methyl 1butanol ns 0.00 0.00 furfuryl acetate ns 0.34 0.20 methyl isobutyrate ns 6.32 4.45 allyl methyl sulfide 0.18 2.26 butyl propionate ns 0.31 0.14 cyclooctene ns 0.60 0.50 isobutyl butyrate ns 0.42 0.20 benzaldehyde ns 0.39 0.93 3phenylpropylacetate ns 0.51 0.38 methyl caprylate ns 1.62 1.64 methyl caproate ns 10.6 7.33 isobutyl propionate ns 1.34 0.91 ethyl 3(methylthio)propionate ns 1.81 2.33 heptyl acetate ns 2.43 2.12 phenethyl acetate ns 1.80 1.76 amyl acetate^ ns 8.19 5.08 methyl butyrate ns 11.8 13.2 cis 6 nonen 1 ol ^ ns 15.0 30.0 isovaleronitrile 1.0 8 0.05 ethyl caproate^ ns 26.7 13.2 cis 3hexenyl acetate ^ ns 18.8 11.8 ben zyl acetate^ 6.45 8.67 ethyl propionate ^ ns 42.3 43.5 ethyl isobutyrate ns 0.00 0.00 isobutyl acetate ^ ns 690 538 propyl acetate^ ns 81.7 104 hexyl acetate ^ ns 78.6 50.6 ethyl butyrate ^ ns 36.8 32.7 butyl acetate ^ ns 94.9 71.6 ethyl 2 methylbutyr ate ^ ns 49.27 25.9 2methylbutyl acetate ^ ns 408 307 methyl 2 methyl butyrate ^ ns 46.9 39.1 Total identified volatiles ns 1646 1322 ^ Sig. contributors ns 1584 1269 ns and *, not significant and significant F test at P

PAGE 168

168 Table B2 2: Stage ZYG means of Galia and Galia type melon aroma compounds, spring 2007. Aroma Compound Signif.z Galia MG10183 propyl butyrate ns 0.92 1.14 tiglic aldehyde ns 0.03 0.01 4methyl 1cyclohexene ns 0.07 0.07 cis 3hexen 1ol ns 0.70 0.16 tr ans cyclodecene ns 0.30 0.32 cinnamyl acetate 0.08 0.20 2methyl 1butanol 0.35 0.07 furfuryl acetate ns 2.23 0.84 methyl isobutyrate ns 0.17 0.13 allyl methyl sulfide 0.12 2.78 butyl propionate ns 0.97 0.50 cyclooctene 1.31 0.52 isobutyl butyrate ns 0.97 0.78 benzaldehyde 0.12 0.03 3phenylpropylacetate 4.43 1.39 methyl caprylate 5.37 14.2 methyl caproate ns 12.9 14.8 isobutyl propionate ns 5.75 3.96 ethyl 3 (methylthio)propionate 2.61 5.70 heptyl acetate ns 7.80 4.93 phenet hyl acetate ns 0.74 0.25 amyl acetate ^ 21.6 15.4 methyl butyrate 6.08 2.38 cis 6nonen1ol ^ 43.0 7.22 isovaleronitrile ns 12.0 0.06 ethyl caproate^ ns 48.8 44.1 cis 3hexenyl acetate ^ ns 35.7 51.2 benzyl acetate ^ 44.3 84.1 ethyl propionate ^ ns 72.6 63.6 ethyl isobutyrate ^ 60.0 0.70 isobutyl acetate ^ 42.1 124.3 propyl acetate^ 156.5 189.9 hexyl acetate^ ns 76.1 172 ethyl butyrate ^ ns 45.1 59.7 butyl acetate^ ns 143 188 ethyl 2methyl butyrate ^ ns 21.9 46.2 2methylbutyl acetate ^ ns 280 313 methyl 2 methyl butyrate ^ 1471 2.7b Total Volatiles ns 2616 1418 ^ Sig. Contributors 2565 1266 ns and *, not significant and significant F test at P

PAGE 169

169 Table B2 3: Stage ZYG means of Galia and Galia type melon aroma compounds, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 1.77 0.91 5.96 tiglic aldehyde 0.22 0.14 0.49 4 methyl 1 cyclohexene 0 .15 0.07 0.27 cis 3 hexen 1 ol 2.32 0.12 2.42 trans cyclodecene 2.49 0.28 0.30 cinnamyl acetate 0.78 0.33 0.32 2 methyl 1 butanol 0.78 0.02 7.88 furfuryl acetate 3.59 0.27 1.45 methyl isobutyrate 0.51 0.22 0.20 allyl methyl sulfide 4.24 3.36 4.74 butyl propionate 2.48 3.98 0.67 3.31 cyclooctene 0.91 0.48 2.71 isobutyl butyrate 3.42 0.92 9.75 benzaldehyde 22.6 2.61 1.00 28.4 3 phenylpropylacetate 4.65 0.49 6.02 methyl caprylate 6.75 7.49 7.77 methyl caproate 14.9 8.78 31.3 isobutyl propionate 14.7 3.59 26.5 ethyl 3 (methylthio)propionate 10.6 8.55 4.55 21.5 heptyl acetate 17.5 9.69 2.59 6.71 phenethyl acetate 33.9 21.6 60.5 amyl acetate 16.6 1.52 6.14 methyl butyrate 23.0 12.1 0.19 32.9 pentyl acetate 27.5 9.96 22.9 cis 6 nonen 1 ol 66.3 20.4 17.6 154 isovaleronitrile 0.24 0.53 0.10 0.18 ethyl caproate 35.3 69.6 19.5 58.3 cis 3 hexenyl acetate 2.50 2.99 0.74 5.13 benzyl acetate 48.9 83.8 67.3 267 ethyl propionate 71.5 6.60 4.49 105 ethyl isobutyrate 13.9 40.6 0.15 21 .6 isobutyl acetate 27.6 137 1.84 44.9 propyl acetate 97.1 144 9.50 232 hexyl acetate 116 20.6 130 251 ethyl butyrate 87.9 31.7 195 butyl acetate 238 74.3 331 ethyl 2 methyl butyrate 29.6 53.7 17.4 73. 4 2 methylbutyl acetate 298 271 320 methyl 2 methyl butyrate 987 6.26 0.7 2433 Total Volatiles 425 1387 715 4790 z Mean separation by Fishers least significant difference test (P

PAGE 170

170 Table B2 4: Stage ZYG means of Galia and Galia type melon aroma compounds following storage for 5 days at 20C, spring 2007. Aroma Compound Signif.z Galia MG10183 propyl butyrate 2.22 1.44 tiglic aldehyde ns 0.03 0.01 4 meth yl 1 cyclohexene ns 0.08 0.03 cis 3 hexen 1 ol ns 0.95 0.63 trans cyclodecene ns 0.19 0.32 cinnamyl acetate ns 0.57 0.33 2 methyl 1 butanol ns 19.2 0.51 furfuryl acetate ns 1.60 1.71 methyl isobutyrate ns 0.04 0.04 allyl methyl sulfide 0.62 1.49 butyl propionate ns 1.74 0.68 cyclooctene ns 0.33 0.69 isobutyl butyrate 1.50 0.69 benzaldehyde ns 0.33 0.08 3phenylpropylacetate 9.75 0.97 methyl caprylate ns 2.65 5.89 methyl caproate 3.12 5.40 isobutyl propionate ns 5.43 2.98 ethyl 3 (met hylthio)propionate ns 6.63 5.74 heptyl acetate ns 7.02 2.68 phenethyl acetate ns 0.13 0.86 amyl acetate^ ns 6.43 4.86 methyl butyrate ns 14.1 7.53 cis 6 nonen 1 ol ns 3.66 5.54 isovaleronitrile ns 3.05 0.05 ethyl caproate^ ns 8.14 10.5 cis 3hexeny l acetate^ ns 71.3 129 benzyl acetate ^ ns 54.3 50.1 ethyl propionate ^ ns 59.4 47.2 ethyl isobutyrate ^ 30.51 9.85 isobutyl acetate ^ ns 71.3 62.7 propyl acetate^ ns 119 148 hexyl acetate^ ns 178 113 ethyl butyrate ^ ns 87.2 69.4 butyl acetate^ ns 142 107 ethyl 2methyl butyrate ^ ns 65.9 33.5 2 methylbutyl acetate ^ ns 289 274 methyl 2 methyl butyrate ^ ns 803 1119 Total Identified volatiles ns 2103 2246 ^ Signif. contributors ns 195 4 2076 ns and *, not significant and significant F test at P respectively.

PAGE 171

171 Table B2 5: Stage ZYG means of Galia and Galia type melon aroma compounds measured after 5 days storage at 20C, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 8.54 12.0 15.7 tiglic aldehyde 0.02 0.0 1 0.03 4methyl 1cyclohexene 0.18 0.17 0.26 cis 3hexen 1ol 3.52 2.06 4.17 trans cyclodecene 0.44 0.71 0.35 cinnamyl acetate 1.42 3.04 0.25 0.45 2 methyl 1 butanol 17.5 60.3 1.92 1.88 furfuryl acetate 4.93 2.96 3.04 methyl isobutyrate 0.07 0. 05 0.12 allyl methyl sulfide 3.47 7.03 4.44 butyl propionate 4.69 5.27 7.46 cyclooctene 0.52 0.67 1.20 isobutyl butyrate 7.00 3.85 7.25 benzaldehyde 4.22 7.60 9.35 23.7 3 phenylpropylacetate 11.9 28.2 3.91 41.3 methyl caprylate 3.53 4.25 3.86 methyl caproate 6.39 6.74 10.4 isobutyl propionate 4.90 11.8 7.67 15.7 ethyl 3 (methylthio)propionate 2.11 5.08 2.18 5.6 1 heptyl acetate 2.50 13.7 4.92 19.2 phenethyl acetate 23.0 19.7 21.3 amyl acetate 9.60 23.5 27.5 methyl butyrate 18.4 39.8 10. 8 27.1 pentyl acetate 51.2 34.7 50.6 cis 6nonen1ol 53.7 25.7 15.2 118.3 isovaleronitrile 3.73 12.6 0.14 0.09 ethyl caproate 22.6 18.3 19.7 cis 3 hexenyl acetate 4.26 2.25 2.39 benzyl acetate 126.3 105 92.8 259 ethyl propionate 28.7 31.5 20.3 ethyl isobutyrate 12.2 3.69 4.92 isobutyl acetate 55.3 120 49.0 33.1 propyl acetate 287 416 246 hexyl acetate 130 578 352 136 ethyl butyrate 44.9 226 294 79.5 butyl acetate 201 187 234 ethyl 2 methyl butyrate 62.4 68.4 64.2 2methylbutyl acetat e 152 551 390 308 methyl 2 methyl butyrate 2356 2368 2564 Total Volatiles 4877 4452 4384 z Mean separation by Fishers least significant difference test (P

PAGE 172

172 Appendix B -3 Table B3 1: Stage HS means of Galia and Galia -type melon aroma compounds, fall 2006. Aroma Compound Signif.z Galia MG10183 propyl butyrate ns 0.28 0.40 tiglic aldehyde ns 0.97 0.80 4 methyl 1 cyclohexene 0.69 1.78 cis 3 hexen 1 ol ns 0.27 0.09 trans cyclodecene ns 0.19 0.10 cinnamyl acetate 0.02 0.08 2 methyl 1 butanol ns 0.01 0.01 furfuryl acetate ns 0.43 0.39 methyl isobutyrate ns 5.21 3.03 allyl methyl sulfide 0.12 3.57 butyl propionate ns 0.19 0.21 cyclooctene ns 0.69 0.49 isobutyl butyrate ns 0.35 0.29 benzaldehyde 0.36 1.10 3 phenylpro pylacetate ns 0.75 0.63 methyl caprylate ns 1.31 1.83 methyl caproate ns 8.21 6.86 isobutyl propionate ns 1.00 1.23 ethyl 3 (methylthio)propionate ns 1.74 1.97 heptyl acetate ns 2.64 2.53 phenethyl acetate ns 3.48 5.45 amyl acetate ^ ns 8.29 7.78 me thyl butyrate ns 11.7 10.3 cis 6 nonen 1 ol ^ ns 19.9 41.8 isovaleronitrile ns 3.05 0.02 ethyl caproate ^ ns 17.0 12.7 cis 3 hexenyl acetate ^ ns 13.6 31.0 benzyl acetate ^ ns 4.43 8.97 ethyl propionate ^ ns 33.16 2 5.5 ethyl isobutyrate ns 0.01 0.01 iso butyl acetate ^ ns 769 702 propyl acetate ^ ns 91.3 101 hexyl acetate ^ ns 73.9 86.1 ethyl butyrate ^ ns 29.9 27.2 butyl acetate ^ ns 95.1 118 ethyl 2 methylbutyrate ^ 29.9 17.1 2 methylbutyl acetate ^ ns 361 405 methyl 2 methyl butyrate ^ ns 37.2 29.7 T otal identified volatiles ns 1624 1656 ^ Sig. contributors ns 1570 1582 ns and *, not signific ant and significant F test at P

PAGE 173

173 Table B3 2: Stage HS means of Galia and Galia -type melon aroma compounds, spring 2007. Aroma Compound Signif. z Galia MG10183 propyl butyrate ns 0.70 1.16 tiglic aldehyde ns 0.25 0.1 0 4 methyl 1 cyclohexene ns 0.07 0.05 cis 3 hexen 1 ol ns 0.35 0.34 trans cyclodecene ns 0.36 0.23 cinnamyl acetate 0.10 0.19 2 methyl 1 butanol ns 0.24 0.23 furfuryl acetate ns 1.14 1.51 methyl isobutyrate ns 0.75 0.42 allyl methyl sulfide ns 0. 09 3.04 butyl propionate 0.96 0.35 cyclooctene ns 0.88 0.53 isobutyl butyrate ns 0.56 0.38 benzaldehyde ns 0.36 0.48 3 phenylpropylacetate ns 2.31 2.22 methyl caprylate ns 2.85 3.11 methyl caproate ns 6.61 6.98 isobutyl propionate ns 4.32 2.21 e thyl 3 (methylthio)propionate ns 3.82 3.93 heptyl acetate ns 4.36 5.89 phenethyl acetate ns 4.30 1.79 amyl acetate ^ ns 13.7 26.0 methyl butyrate ns 14.3 3.87 cis 6 nonen 1 ol ^ ns 5.07 13.0 isovaleronitrile 13.5 0.08 ethyl caproate ^ ns 23.9 23.7 c is 3 hexenyl acetate 20.6 142 benzyl acetate ^ 18.8 95.0 ethyl propionate ^ ns 42. 75.4 ethyl isobutyrate ^ ns 21.8 35.6 isobutyl acetate ^ ns 26.7 53.0 propyl acetate ^ ns 124 206 hexyl acetate ^ ns 106 214 ethyl butyrate ^ ns 46.8 39.3 butyl acetate ^ ns 135 152 ethyl 2 methyl butyrate ^ ns 33.3 102 2 methylbutyl acetate ^ ns 173 234 methyl 2 methyl butyrate ^ ns 1074 1184 Total Volatiles ns 1914 2631 ^ Sig. Contributors ns 1833 2384 ns and *, not significant and significant F test at P tively.

PAGE 174

174 Table B3 3: Stage HS means of Galia and Galia -type melon aroma compounds, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 2.18 1.84 6.08 tiglic aldehyde 0.01 0.06 0.09 4 methyl 1 cyclohexene 0.16 0.0 5 0.26 cis 3 hexen 1 ol 0.89 3.20 3.93 trans cyclodecene 0.50 0.27 0.47 cinnamyl acetate 0.43 0.23 1.48 2 methyl 1 butanol 0.39 0.13 2.17 furfuryl acetate 1.79 1.37 2.69 methyl isobutyrate 0.10 0.17 0.24 allyl methyl sulfide 2.06 10.0 7.60 b utyl propionate 2.49 3.44 6.70 cyclooctene 0.90 0.46 2.99 isobutyl butyrate 2.09 1.15 7.24 benzaldehyde 1.36 3.84 15.2 3 phenylpropylacetate 4.35 3.20 42.4 methyl caprylate 4.56 5.12 11. 7 methyl caproate 13.8 9.33 19.5 isobutyl propionate 9. 56 5.18 31.2 ethyl 3 (methylthio)propionate 3.54 5.29 7.02 heptyl acetate 10.1 6.88 31.7 phenethyl acetate 12.7 37.0 63.2 amyl acetate 56.9 28.4 69.3 methyl butyrate 15.1 12.5 20. pentyl acetate 40.0 24.2 54.3 cis 6 nonen 1 ol 43.5 4.83 20.8 8 9.1 isovaleronitrile 1.23 0.04 0.18 ethyl caproate 35.5 21.2 36.1 cis 3 hexenyl acetate 2.18 2.24 9.08 benzyl acetate 85.4 125 286 ethyl propionate 17.9 37.6 53.2 ethyl isobutyrate 23.4 6.07 27.1 isobutyl acetate 49.1 147 27.1 99.7 propyl ace tate 175 19.5 322 hexyl acetate 177 276 428 ethyl butyrate 62.1 86.5 99.5 butyl acetate 215 86.0 214 ethyl 2 methyl butyrate 65.8 37.1 67.2 2 methylbutyl acetate 205 384 106 3 84 methyl 2 methyl butyrate 1811 4.15 1.4 1854 Total Volatiles 3297 1 545 1007 5702 z Mean separation by Fishers least significant difference test (P

PAGE 175

175 Table B3 4: Stage HS means of Galia and Galia type melon aroma compounds following storage for 3 days at 20C, spring 2007. Aroma Compound Signif. z Galia MG10183 propyl butyrate ns 1.59 2.37 tiglic aldehyde ns 0.06 0.02 4 methyl 1 cyclohexene ns 0.05 0.10 cis 3 hexen 1 ol ns 0.60 0.83 trans cyclodecene ns 0.16 0.28 cinnamyl acetate ns 0.90 0.30 2 methyl 1 butanol ns 3.96 1.11 furfuryl aceta te ns 0.88 1.48 methyl isobutyrate ns 0.52 0.04 allyl methyl sulfide ns 0.36 2.23 butyl propionate ns 1.19 1.43 cyclooctene ns 2.32 0.48 isobutyl butyrate ns 1.37 1.21 benzaldehyde ns 0.93 0.10 3 phenylpropylacetate ns 4.72 3.73 methyl caprylate ns 4.73 4.99 methyl caproate ns 5.36 6.84 isobutyl propionate ns 5.31 3.94 ethyl 3 (methylthio)propionate ns 8.44 3.58 heptyl acetate ns 5.43 9.45 phenethyl acetate ns 1.48 0.19 amyl acetate ^ ns 13.1 16.3 methyl butyrate ns 13.4 8.98 cis 6 nonen 1 ol ns 3.79 4.46 isovaleronitrile ns 12.5 0.25 ethyl caproate ^ ns 19.6 16.2 cis 3 hexenyl acetate ^ ns 32.9 125 benzyl acetate ^ ns 55.9 69.7 ethyl propionate ^ ns 38.2 46.5 ethyl isobutyrate ^ ns 16.2 19.8 isobutyl acetate ^ ns 53.5 47.9 propyl acetate ^ n s 135 121 hexyl acetate ^ ns 159 166 ethyl butyrate ^ ns 98.9 59.1 butyl acetate ^ ns 120 126 ethyl 2 methyl butyrate ^ ns 42.7 36.5 2 methylbutyl acetate ^ ns 261 297 methyl 2 methyl butyrate ^ ns 1167 1173 Total Identified volatiles ns 2294 2395 ^ Sign if. contributors ns 2198 2216 ns and *, not significant and significant F test at P

PAGE 176

176 Table B3 5: Stage HS means of Galia and Galia type melon aroma compounds measured after 3 days storage at 20C, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 6.32 5.65 8.39 tiglic aldehy de 0.05 0.10 0.02 4 methyl 1 cyclohexene 0.32 0.17 0.11 cis 3 hexen 1 ol 0.67 1.68 1.00 2.37 trans cyclodecene 0.50 0.45 0.23 cinnamyl acetate 0.41 1.06 0.14 0.74 2 methyl 1 butanol 2.72 1.15 6.37 furfuryl acetate 2.37 1.63 1.64 methyl isobutyr ate 0.08 0.03 0.03 allyl methyl sulfide 2.52 2.59 8.51 5.65 butyl propionate 3.53 3.38 5.04 cyclooctene 0.76 0.42 1.30 isobutyl butyrate 4.18 2.16 6.24 benzaldehyde 7.35 3.78 6.39 17.1 3 phenylpropylacetate 6.98 9.70 5.00 34.9 methyl caprylate 3.97 3.78 10.1 methyl caproate 9.39 11.0 19.8 isobutyl propionate 3.20 8.96 7.62 13.7 ethyl 3 (methylthio)propionate 4.26 3.42 7.19 heptyl acetate 7.45 13.2 8.07 25.2 phenethyl acetate 17.3 37.2 39.4 amyl acetate 13.8 13.3 17.1 methyl butyrate 34.3 22.8 23. pentyl acetate 39.4 35.9 45.2 cis 6 nonen 1 ol 20.5 15.7 15.0 69.3 isovaleronitrile 0.56 0.55 0.11 ethyl caproate 38.6 28.9 38.8 cis 3 hexenyl acetate 3.65 3.49 7.55 benzyl acetate 102 82.7 73.2 315 ethyl propionate 45.5 35.8 116 2 9.9 ethyl isobutyrate 9.35 16.68 16.55 isobutyl acetate 199 43.1 71.3 propyl acetate 222 218 239 hexyl acetate 501 426 463 ethyl butyrate 143 78.2 121 butyl acetate 426 90.3 173 ethyl 2 methyl butyrate 60.7 94.9 62.2 2 methylbutyl acetate 4 62 218 280 methyl 2 methyl butyrate 1374 1608 2262 Total Volatiles 3756 3208 4440 z Mean separation by Fishers least significant difference test (P

PAGE 177

177 Appendix B -4 Table B4 1: Stage FS means of wild-type (WT) Galia and Galia type melon aroma compounds, fall 2006. Aroma Compound Signif.z Galia MG10183 propyl butyrat e ns 0.63 0.45 tiglic aldehyde ns 1.80 1.11 4 methyl 1 cyclohexene ns 0.80 1.33 cis 3 hexen 1 ol 0.50 0.09 trans cyclodecene 0.16 0.11 cinnamyl acetate ns 0.08 0.07 2 methyl 1 butanol ns 0.007 0.01 furfuryl acetate ns 0.65 0.66 methyl isobutyra te 5.39 3.35 allyl methyl sulfide ns 1.32 4.23 butyl propionate ns 0.63 0.26 cyclooctene ns 0.94 0.75 isobutyl butyrate ns 0.51 0.24 benzaldehyde 0.54 1.33 3 phenylpropylacetate ns 1.46 0.97 methyl caprylate ns 1.78 1.58 methyl caproate ns 7.26 7.46 isobutyl propionate ns 1.75 1.10 ethyl 3 (methylthio)propionate ns 2.22 1.37 heptyl acetate ns 4.29 3.67 phenethyl acetate ns 3.79 9.20 amyl acetate ^ ns 12.1 11.9 methyl butyrate ns 14.4 13.4 cis 6 nonen 1 ol ^ ns 31.9 64.1 isovaleronitrile 8.21 0.15 ethyl caproate ^ ns 20.4 14.4 cis 3 hexenyl acetate ^ ns 35.2 55.6 benzyl acetate ^ 5.95 11.0 ethyl propionate ^ ns 33.9 30.6 ethyl isobutyrate ns 0.03 0.01 isobutyl acetate ^ ns 939 949 propyl acetate ^ 106 163 hexyl acetate ^ ns 121 144 e thyl butyrate ^ ns 39.5 33.4 butyl acetate ^ ns 194 220 ethyl 2 methylbutyrate ^ 35.3 20.1 2 methylbutyl acetate ^ ns 519 635 methyl 2 methyl butyrate ^ ns 45.8 42.1 Total identified volatiles ns 2190 2446 ^ Sig. contributors ns 2104 2337 ns and *, not significant and significant F test at P

PAGE 178

178 Table B4 2: Stage FS means of Galia and Galia type melon aroma compounds, Spring 2007. Aroma Compound Signif. z Galia MG10183 propyl butyrate ns 0.75 0.68 tiglic aldehyde ns 0.43 0.32 4 methyl 1 cyclohexene ns 0.03 0 .03 cis 3 hexen 1 ol ns 0.36 0.19 trans cyclodecene ns 0.76 0.40 cinnamyl acetate ns 0.16 0.13 2 methyl 1 butanol ns 1.06 0.43 furfuryl acetate 1.27 1.60 methyl isobutyrate ns 1.47 1.27 allyl methyl sulfide ns 0.21 1.46 butyl propionate 0.98 0. 34 cyclooctene ns 2.61 1.50 isobutyl butyrate ns 1.21 0.38 benzaldehyde ns 1.36 1.14 3 phenylpropylacetate ns 2.78 1.88 methyl caprylate ns 2.42 3.54 methyl caproate ns 8.40 6.44 isobutyl propionate ns 3.33 2.07 ethyl 3 (methylthio)propionate 4.0 4 2.24 heptyl acetate ns 7.17 5.30 phenethyl acetate ns 4.47 4.66 amyl acetate ^ ns 16.9 22.0 methyl butyrate ns 15.2 8.43 cis 6 nonen 1 ol ^ ns 7.57 6.72 I sovaleronitrile ^ 6.31 0.06 ethyl caproate ^ ns 33.4 19.6 cis 3 hexenyl acetate ^ 20.1 84.4 benzyl acetate ^ 29.0 91.3 ethyl propionate ^ ns 50.0 70.7 ethyl isobutyrate ^ ns 51.7 33.2 isobutyl acetate ^ ns 24.5 51.0 propyl acetate ^ ns 49.9 129 hexyl acetate ^ ns 84.8 104 ethyl butyrate ^ ns 51.5 52.0 butyl acetate ^ ns 84.1 122 ethyl 2 methyl butyrate ^ ns 66.7 62.6 2 methylbutyl acetate ^ ns 130 90.9 methyl 2 methyl butyrate ^ ns 273 482 Total Volatiles ns 1034 1464 ^ Sig. Contributors ns 938 1306 ns and *, not significant and significant F test at P

PAGE 179

179 Table B4 3: Stage FS means of Galia and Galia type muskmelon aroma compounds, fall 2007. Aroma Compound LSD z Galia MG10183 Elario propyl butyrate 1.8 1.02 2.34 3.58 tiglic aldehyde 0.10 0.12 0.09 4 methyl 1 cyclohexene 0.18 0.29 0.07 cis 3hexen 1ol 0.58 0.56 0.81 trans cyclodecene 0.21 0.51 0.23 cinnamyl acetate 0.39 0.26 0.41 1.72 2 methyl 1 butanol 0.39 0.29 1.09 furfuryl acetate 0.99 1.53 1.59 methyl isobutyrate 0.07 0.25 0.05 allyl methyl sulfide 1.54 7.01 4.25 butyl propionate 1 .34 1.63 3.21 cyclooctene 0.54 0.41 0.79 isobutyl butyrate 1.20 1.39 2.33 benzaldehyde 1.16 6.45 6.82 3phenylpropylacetate 17.4 2.84 4.09 25.1 methyl caprylate 2.09 4.96 4.58 methyl caproate 5.84 14.5 12.3 isobutyl propionate 7.02 6.43 7.65 ethyl 3(methylthio)propionate 2.67 3.14 3.87 heptyl acetate 7.73 7.17 9.43 20.8 phenethyl acetate 14.4 40.4 23.6 amyl acetate 57.0 38.1 35.8 methyl butyrate 12.4 5.24 0.64 cis 6nonen1ol 16.9 5.73 53.7 59.7 isovaleronitrile 1.83 4.44a 0.08b 0. 11b ethyl caproate 29.1 48.3 34.9 cis 3 hexenyl acetate 2.46 1.30 4.28 4.88 benzyl acetate 87.9 141 168 ethyl propionate 12.0 29.7 14.7 ethyl isobutyrate 22.8 46.5 1.98 24.2 isobutyl acetate 183 44.8 84.6 propyl acetate 170 239 150 hexyl acetate 114 168 336 304 ethyl butyrate 94.2 58.5 81.5 butyl acetate 266 235 138 ethyl 2 methyl butyrate 68.7 44.1 1.32 2 methylbutyl acetate 327 320 214. methyl 2 methyl butyrate 270.7 34.5 3.68 1199 Total Volatiles 1616 1710 2700 Signif. Contributo rs 798.8 1542 1568 25 59 z Mean separation by Fishers least significant difference test (P

PAGE 180

180 Table B4 4: Stage FS means of Galia and Galia type melon aroma compounds following storage for 2 days at 20C, spring 2007. Aroma Compound Signif z Galia MG10183 propyl butyrate ns 2.40 1.55 tiglic aldehyde ns 0.04 0.04 4 methyl 1 cyclohexene ns 0.03 0.02 cis 3 hexen 1 ol ns 0.72 0.23 trans cyclodecene ns 0.40 0.32 cinnamyl acetate ns 0.32 0.27 2 methyl 1 butanol ns 1.43 0.64 furfuryl a cetate ns 1.43 1.10 methyl isobutyrate ns 0.37 1.23 allyl methyl sulfide ns 0.17 3.00 butyl propionate 2.38 1.10 cyclooctene ns 1.83 1.44 isobutyl butyrate ns 1.61 0.83 benzaldehyde ns 0.78 1.87 3 phenylpropylacetate 5.21 2.33 methyl caprylate ns 2.57 2.84 methyl caproate ns 7.55 8.00 isobutyl propionate ns 6.35 4.11 ethyl 3 (methylthio)propionate ns 6.17 3.07 heptyl acetate ns 10.4 7.33 phenethyl acetate ns 1.75 3.57 amyl acetate ^ ns 20.0 15.8 methyl butyrate ns 16.1 12.2 cis 6 nonen 1 ol ns 5.14 4.90 I sovaleronitrile ^ ns 24.3 0.72 ethyl caproate ^ ns 22.8 22.2 cis 3 hexenyl acetate ^ ns 77.2 176 benzyl acetate ^ ns 46.2 66.3 ethyl propionate ^ ns 39.7 37.1 ethyl isobutyrate ^ ns 41.5 25.2 isobutyl acetate ^ ns 36.5 52.7 propyl acetate ^ ns 156 159 hexyl acetate ^ ns 173 87.4 ethyl butyrate ^ ns 103 53.0 butyl acetate ^ ns 79.1 97.9 ethyl 2 methyl butyrate ^ ns 56.4 50.6 2 methylbutyl acetate ^ ns 196 206 methyl 2 methyl butyrate ^ ns 1188 1701 Total Identified volatiles ns 2330 2828 ^ Signif. contributors ns 2183 2596 ns and *, not significant and significant F test at P

PAGE 181

181 Table B4 5: Stage FS means of Galia and Galia type melon aroma compounds measured after 2 days storage at 20C, fall 2007. Aroma Compound LSDz Galia MG10183 Elario propyl butyrate 5.25 3.06 6.16 tiglic aldehyde 0.01 0.25 0.01 4methyl 1cyclohexene 0.18 0.13 0.27 cis 3 hexen 1 ol 1.27 1.02 0.82 2.85 trans cyclodecene 0.35 0.55 0.44 cinnamyl acetate 0.99 0.98 0.20 2.17 2 methyl 1 butanol 0.30 0.19 0.44 furfuryl acetate 2.27 1.52 2.41 methyl isobutyra te 0.06 0.02 0.10 0.17 allyl methyl sulfide 2.81 8.66 3.41 butyl propionate 4.16 2.39 3.87 cyclooctene 0.81 0.72 0.30 isobutyl butyrate 3.46 1.46 3.16 benzaldehyde 4.33 2.9 4.99 9.99 3 phenylpropylacetate 6.76 7.94 5.90 33 .5 methyl caprylate 4. 31 7.24 4.35 methyl caproate 10.3 9.97 13.6 isobutyl propionate 4.22 7.38 6.03 13.3 ethyl 3 (methylthio)propionate 1.20 4.52 3.02b 4.72 heptyl acetate 12.9 20.2 13.6 33.1 phenethyl acetate 32.0 32.2 24.3 amyl acetate 22.8 9.97 8.53 methyl butyrat e 26.3 28.4 12.8 0.20 pentyl acetate 27.8 30.8 31.9 cis 6 nonen 1 ol 7.09 42.4 36.9 isovaleronitrile 3.77 0.13 0.05 ethyl caproate 36.5 24.1 31.3 cis 3 hexenyl acetate 3.76 3.21 7.36 benzyl acetate 63.3 104 202 ethyl propionate 8.73 12.0 24.4 ethyl isobutyrate 1.51 10.4 2.44 0.94 isobutyl acetate 63.8 57.6 48.7 propyl acetate 234 159 215 hexyl acetate 486 470 168 ethyl butyrate 116 95.7 350 butyl acetate ns 127 265 189 ethyl 2 methyl butyrate ns 59.0 35.0 55.4 2 methylbutyl acetate ns 392 245 222 methyl 2 methyl butyrate 913 848 1309 2067 Total Volatiles 866 2645 2981 3822 z Mean separation by Fishers least significant difference test (P

PAGE 182

182 Chapter 4 ANOVA Tables Table 4 2, fall 2006. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 15.1* 0.1 360 5.61 23.1 2 0.0002 0.01 15.6 2.28 15.6 Rep 3 3.45 0.06 121 50.6 22.5 1.08 0.0001 0.00005 30.6 1.12 30.6 Error 3 1.45 0.17 420 232.2 18.3 0.75 0.0001 0.04 9.04 1.11 9.04 *, significant F test at P Table 4 2, spring, 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 648** 0.45* 2145.1* 512* 26.6 1.28 0.04 0.0001 331.5* 0.21 83.9 Rep 3 16.5 0.03 56.1 207 8.01 2.3 0.02 0.0001 37.2 0.02 26.5 Error 3 13.3 0 .03 172.8 41.7 20.2 0.56 0.01 0.0001 11.9 0.03 38.3 and **, significant F test at P Table 4 2, fall, 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 2 4.11 0.13* 117.5 413.8* 33.2 4.75 0.07 0.00001 94.7 5.78* 19 Rep 2 0.78 0.08 93.8 180.2 12.3 0.78 0.004 0.00001 32.2 0.04 82.1 Error 4 2.11 0.01 22.6 37.9 2.27 1.53 0.05 0.00001 25.5 0.33 87.7 *, significant F test at P Table 4 3. TIV, fall 2006. Table 4 3. TIV, spring 2007.

PAGE 183

183 At Harvest At Harvest After St orage Source DF Mean square Source DF Mean square DF Mean square Trt 3 74249 Trt 1 15.8 1 5408813 Rep 1 309861 Rep 3 173582 3 138839 Error 3 56086 Error 3 63160 3 11056 Table 4 3. TIV, fall 2007. At Harvest After Storage Source DF Mean square DF Mean square Trt 2 460095 2* 182146 Rep 2 220817 2 15346 Error 4 148963 4 13855 Table 4 4 fall 2006. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 21.1* 0.13 544.5 98 4.06 0.91 0 .00 0.05 103** 99.8* 220.6 Rep 3 1.46 0.03 58.8 44.3 28.2 0.68 0.0004 0.03 9.55 1.28 67 Error 3 1.46 0.08 86.8 100.3 23 1.71 0.00003 0.01 1.86 2.93 56.5 and **, significant F test at P ly.

PAGE 184

184 Table 4 4 ., spring 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 72* 0.25* 979* 420.5* 12.8 1.53 0.11* 0.001 1369** 10.6 91.1 Rep 3 3.5 0.04 111.2 100.5 32.5 0.61 0.01 0.0001 0.55 4.5 41.4 Error 3 4.33 0.01 59.9 25.8 4.66 0.53 0.01 0.0002 3.27 3.1 2.44 and **, significant F test at P Table 4 4., fall 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH TA Firmness Ethylene Respiration Trt 2 10.1* 0.31 353.2 438.4 91.8 3.92 0.01 0.0008** 251.8 5.78* 11.8 Rep 2 0.7 8 0.04 48.1 89.9 12.9 0.68 0.02 0.00001 83.1 0.04 14.6 Error 4 1.28 0.08 191.6 82.9 28.6 1.19 0.02 0.00004 38.4 0.33 35.8 *and **, significant F test at P Table 4 5. TIV, fall 2006. Table 4 5. TIV, spring 2007. At H arvest At Harvest After Storage Source DF Mean square Source DF Mean square DF Mean square Trt 3 209217 Trt 1 2871429 1 40792 Rep 1 125334 Rep 3 170127 3 90822 Error 3 292381 Error 3 292802 3 177041

PAGE 185

185 Table 4 5. TIV, fall 2007. At Harvest After Storage Source DF Mean square DF Mean square Trt ** 2 14314161 2 2135686 Rep 2 445716 2 774529 Error 4 680519 4 65565 **, significant F test at P Table 4 6, fall 2006. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 66.1** 0.08 84.5 215.3* 10.6 0.78 0.0000 0.00001 76.6 184.4* 10.4 Rep 3 8.79 0.13 130.3 137.1 2 3.7 1.53 0.0002 0.01035 5.32 26.8 14.2 Error 3 1.13 0.04 64.6 13.6 42.5 1.78 0.00003 0.00301 12.4 16.3 5.04 *and **, significant F test at P Table 4 6, spring 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 60.5** 0.18 780.1* 318.8* 49.0 3.78 0.002 0.0002 548.6** 30.4 7.8 Rep 3 5 .0 0.003 24.5 13.3 1.54 0.6 0.008 0.0002 1.04 2.22 2.27 Error 3 0.17 0.02 52.5 24.1 13.6 0.08 0.003 0.00003 2.7 4.87 4.19 *and **, significant F test at P

PAGE 186

186 Table 4 6, fall 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH TA Firmness Ethylene Respiration Trt 2 5.44* 0.39 258.5 615.1 54.1 3.48* 0.03 0.0004 134.8* 5.67 25.6* Rep 2 0.1 1 0.01 3.94 17.2 8.34 0.11 0.05 0.0002 24.2 0.15 5.89 Error 4 0.28 0.07 161.1 88 13.4 0.21 0.15 0.0003 8.12 1.24 1.43 *, significant F test at P Table 4 7. TIV, fall 2006. Table 4 7. TIV, spring 2007. At Harvest At Harvest After Storag e Source DF Mean square Source DF Mean square DF Mean square Trt 3 2068 Trt 1 1028317 1 20423 Rep 1 218821 Rep 3 52498 3 278739 Error 3 155854 Error 3 521031 3 271895 Table 4 7. TIV, fall 2007. At Harvest After Storage Source DF Mea n square DF Mean square Trt* 2 19804441 2 1141930 Rep 2 252598 2 197615 Error 4 2115017 4 510556 *, significant F test at P

PAGE 187

187 Table 4 8 TIV, fall 200 6 Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 1.13 0.13 214.2 2.65* 0.28 6.3 0.0002 0.002 68.4 10.1 1.2 Rep 3 0.46 0.03 72.1 32 3.58 0.64 0.00008 0.01 7.77 7.99 2.22 Error 3 0.13 0.09 59.3 69.2 11.9 0.37 0.001 0.01 22 10.9 9.87 *, significant F test at P Table 4 8 TIV, spring 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH T T A Firmness Ethylene Respiration Trt 1 12.5** 0.18 743.1** 268* 34.9 4.81 0.00001 0.0006 267.4** 4.35 0.05 Rep 3 0.33 0.01 33.2 22.3 14 0.06 0.002 0.0001 3.89 1.07 3.86 Error 3 0.17 0.003 6.76 8.49 7.3 0.27 0.003 0.0001 3.3 0.68 1.19 *and **, significant F test at P Table 4 8 TIV, fall 2007. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC pH TA Firmness Ethylene Respiration Trt 2 4** 0.12** 141.3** 292.9* 18.8 3.63* 0.06 0.002** 68.4* 14.9 4.91 R ep 2 1 0.03 38.1 17.3 0.44 0.15 0.02 0.001 0.02 3.22 4.74 Error 4 0 0.01 4.51 39.5 6.04 0.33 0.02 0.0001 9.09 2.24 11.1 *and **, significant F test at P

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188 Table 4 9. TIV, fall 2006. Table 4 9. TIV, spring 2007. At Har vest At Harvest After Storage Source DF Mean square Source DF Mean square DF Mean square Trt 3 130812 Trt 1 370536 1 497379 Rep 1 286230 Rep 3 285832 3 79320 Error 3 29992 Error 3 142306 3 1311701 Table 4 9. TIV, fall 2007. At H arvest After Storage Source DF Mean square DF Mean square Trt 2 1081594 2 1102426 Rep 2 213342 2 153950 Error 4 157529 4 155945 Table 4 11, spring 2008. Source DF Mean square SSC Firmness Methyl isobutyrate Allyl methyl sulfide ethyl propionate Propyl acetate Methyl butyrate Isovaleronitrile 2 methyl 1 butanol Trt 3 6.57** 524** 24.3* 0.18 2239 13.1 59 148** 110** Rep 2 0.08 13.8 1.36 0.06 485 14.5 21.7 6.88 0.42 Error 6 0.21 4.34 4.54 0.33 534 6.35 13.7 6.96 1.99 *and **, significant F test at P

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189 Table 4 11, spring 2008. Source DF Mean square Tiglic aldehyde 4 -methyl 1 cyclohexene Ethyl isobutyrate Isobutyl acetate Methyl 2 methyl butyrate Ethyl butyrate Butyl acetate Ethyl 2 methyl butyrate Cis 3 hexen 1 ol Trt 3 2.13** 0.002 5213** 347 1495818** 628 1148 2292 1.71** Rep 2 0.14 0.002 223 89.4 9062 38.2 2.22 1651 0.17 Error 6 0.11 0.001 115 23.4 41438 75.1 386 786 0.16 *and **, significant F test a t P Table 4 11, spring 2008. Source DF Mean square Isobutyl propionate 2 methyl butyl acetate Propyl butyrate Butyl propionate Amyl acetate Cyclooctene Methyl caproate Isobutyl butyrate Benzaldehyde Trt 3 9.08* 6078 2.59** 2.08* 569** 0.17 9.17 3.28** 13.8* Rep 2 1.38 4388 0.16 0.05 51.6 0.15 5.7 0. 38 0.16 Error 6 1.47 3662 0.17 0.31 46.5 0.11 2.68 0.22 1.45 *and **, significant F test at P v ely. Table 4 11, spring 2008. Source DF Mean square Ethyl caproate Cis 3 hexenyl acetate Hexyl acetate Ethyl 3 (methylthio) propionate Heptyl acetate Methyl caprylate Trans cyclodecene Cis 6 n onen 1 ol 3 phenyl propylacetate Trt 3 2834** 2078* 218 3.93** 490** 29.6 0.15** 11220** 366** Rep 2 123 213 108 0.17 22.1 7.83 0.001 732 7.16 Error 6 106 375 327 0.14 13.1 8.43 0.002 832 4.56 *and **, significant F test at P v ely.

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190 Table 4 11, spring 2008. Source DF Mean square Benzyl acetate Phenethyl acetate Cinnamyl acetate Total volatiles Signif. Contribs. Trt 3 2402 139** 0.05* 1902374** 1681572 Rep 2 1356 1.37 0.01 7736 5165 Error 6 820 1.64 0.01 58115 59794 *an d **, significant F test at P v ely.

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191 APPENDIX C CHAPTER 5 ANOVA TABLES

PAGE 192

192 Table C5 2. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC TTA pH Firmness Ethylene Respiration Line 3 40.3* 0.22* 416* 260* 60.8* 11.6* 0.01* 0.12 11.2 0.2 46.3 Seas 2 1919** 0.88** 1911** 1412** 240** 38.4** 0.001** 0.26** 34.0 4.57** 239** L*S 6 126 0.03 55.1 23.1 18.8 3.07 0.001 0.08 18.0 0.44 25.4 Seas(rep) 6 219 0.06 39.2 42.3 7.2 2.27 0.001 0.1 26.8 0.13 60.5 Error 18 8.39 0.06 95.7 53.4 7.35 1.18 0.0002 0.06 55.2 0.25 42.7 *and **, significant F test at P v ely. Table C5 3. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC TTA pH Firmness Ethylene Respiration Line 3 38.4** 0.04 54.9 89.2 14.4 1.38** 0.001** 0.44** 449** 37.3** 36.9 Seas 2 352* 0.71** 1461** 1387** 46.6* 23.1 0.001* 0.29** 407** 7.7 154** L*S 6 1.93 0.08 164.6 60 14.6 1.84 0.001** 0.16** 5.8 16.8* 88.7** Seas(rep) 6 2.53 0.05 97.8 29.7 7.63 1.17 0.001 0.03 20.2 5.94 26.3 Error 18 2.75 0.04 76.4 41.8 10 .1 1.06 0.0001 0.01 14.3 4.36 21.9 *and **, significant F test at P v ely.

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193 Table C5 5. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC TTA pH Firmness Ethylene Respiration Line 3 26.1** 0.01 43.2 10 8.18 2.32 0.0002 0.2** 93.6** 28.2** 22.8 Seas 2 342** 1.33** 1868** 2077** 170** 15.8** 0.001** 0.4** 174** 34.1** 132** L*S 6 7.03 0.1 178 102 37.2 2.96 0.001** 0.14** 17.5 18.5** 46.6 Seas(rep) 6 6.14 0.05 53.9 38.7 2.83 0.41 0.0001 0.03 14.8 1.97 14.1 Error 18 3.47 0.05 79 48. 5 18.9 1.15 0.0002 0.02 9.1 2.73 23.1 *and **, significant F test at P v ely. Table C5 7. Source DF Mean square Days to harvest Weight Length Width Flesh thickness SSC TTA pH Firmness Ethylene Respiration Line 3 35** 0.19** 351** 111* 32.9* 11.9** 0.001 0.07 46.3** 14.6 25.5 Seas 2 335** 0.91** 1518** 1449** 97.3** 25.9** 0.0005** 0.28* 86.8* 64.9** 58.6** L*S 6 4.64* 0.06* 149* 73.2 7.76 1.56 0.002 0.06 6.32 9.49 10.1 Seas(rep) 6 1.58 0.005 9.55 12.6 2.58 0.81 0.0003 0.05 10.3 1.62 5.58 Error 18 1.62 0.02 50.4 33.1 8.53 1.37 0.0002 0.05 4.68 7.25 7.71 *and **, significant F test at P v ely.

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194 Table C5 9. Source DF Mean square Stage ZG TIV at Harvest. Stage ZYG TIV at Harvest Stage HS TIV at Harvest. Stage FS TIV at Harvest. Line 3 6198 1761672** 1621190 363025 Seas 2 5 2671 1285478* 2033139 6685365* L*S 6 50924 1092316** 1134116 325884 Seas(rep) 6 22444 162349 625156 796406 Error 18 31354 122644 615920 263086 *and **, significant F test at P v ely. Table C5 10. Source DF Mean square Stage ZG TIV after Storage, Fa07 Stage ZYG TIV after Storage, Fa07. Stage HS TIV after Storage, Fa07. Stage FS TIV after Storage, Fa07. Rep 2 1104 81327 49234 55406 Line* 2 1715599* 8894705** 7273443** 4800206** Error 4 72464 238363 238152 180146 and **, significant F test at P v ely.

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BIOGRAPHICAL SKETCH Jeanmarie Mink Harty (ne Mitchell) was born in 1975 to, Edward A. and Eileen J. Mitchell. R aised in Collegeville, PA, Jeanmarie was active throughout elementary and high school in her local 4H Club, which sparked her interest in agriculture. She received her Bachelor of Science degree in turfgrass s cience from the Pennsylvania State University in 1997. Instead of becoming a golf course superintendent, she traveled throughout Europe in fall 1997 and join ed the U.S. Peace Corps in May 1998. As a Schools Self -Reliance Project Officer in the Kingdom of Lesotho, Southern Africa, Jeanmarie worked to ensure food-security at local primary schools. After successfully completing Peace Corps in June 2000, Jeanmaries love for Lesotho led her to remain in the country for a third year and work as the Agriculture Director of a local NGO called GROW. Whil e at GROW Jeanmarie assisted and educated farmers in sustainable agricultural practices and worked to improve the Basothos livelihoods. In 2001, Jeanmarie left Lesotho and moved to St. Croix, USVI where she was a Research Analyst II at the Agricultural Experiment Station of the University of the Virgin Islands (UVI). At UVI, Jeanmarie conducted research on sustainable agricultural practices. While on St. Croix, Jeanmarie met her husband, Cheyenne. In fall, 2003, Jeanmarie and Cheyenne moved to Gainesv ille, Florida where Jeanmarie was accepted to graduate s chool at the University of Florida, Department of Horticultural Sciences. Under the superior guidance of Daniel J. Cantliffe, Jeanmarie successf ully learned how to grow melons, conduct research and meet the requirements of a Ph.D. Upon graduation, it is the goal of Jeanmarie to continue to work in agriculture with a focus on improving production, quality, and postharvest practices for growers world -wide.