- र र -१ -२ Studies on effects of sugary-1 and shrunken-2 genes on grain sweetness in sweet corn genotypes BRIJESH KUMAR MEHTA DIVISION OF GENETICS ICAR-INDIAN AGRICULTURAL RESEARCH INSTITUTE NEW DELHI – 110 012 2015 Studies on effects of sugary-1 and shrunken-2 genes on grain sweetness in sweet corn genotypes by BRIJESH KUMAR MEHTA A Thesis Submitted to the Faculty of Post Graduate School, ICAR-Indian Agricultural Research Institute, New Delhi, in partial fulfillment of the requirements for the award of the degree of MASTER OF SCIENCE in GENETICS 2015 Approved by a Chairman : (Dr. Firoz Hossain) Co-chairman : (Dr. T. Nepolean) Member : (Dr. M. Vignesh) Member : (Dr. S. K. Jha) Member : (Dr. Amol Kumar Solanke) Member : (Dr. Supradip Saha) Division of Genetics ICAR-Indian Agricultural Research Institute New Delhi – 110012, India Dr. FIROZ HOSSAIN Phone: +91-11-25841285 Senior Scientist e-mail: [email protected] CERTIFICATE This is to certify that the thesis entitled “Studies on effects of sugary-1 and shrunken-2 genes on grain sweetness in sweet corn genotypes” submitted to the Faculty of the Post-Graduate School, ICAR-Indian Agricultural Research Institute, New Delhi, in partial fulfilment of the requirements for the award of Master of Science in GENETICS, embodies the results of bonafide research work carried out by Mr. Brijesh Kumar Mehta, Roll No.: 20425 under my guidance and supervision, and that no part of this thesis has been submitted for any other degree or diploma. It is further certified that any help or source of information that has been availed of in this connection has been duly acknowledged by him. Date : 29.07.2015 (Dr. Firoz Hossain) Place: New Delhi Chairman (Advisory Committee) Aknowledgements It is by the lavish and boundless blessing of the Almighty that I have been able to complete my studies successfully hitherto and present this humble piece of work, for which I am eternally indebted. Fervently and modestly, I extol the genuine cooperation, inspiration and affection offered to me by the Chairman of advisory committee, Dr. Firoz Hossain, Senior Scientist,Division of Genetics right from the initiation of my work to ship-shaping of the manuscript. The present work bears at every stage the impression of his concrete suggestions, careful, seasoned criticism, indefatigable guidance and meticulous attention to details. He has always been a fountain of inspiration to me. It was indeed a rare privilege for me to work under his emending inspiration and indomitable spirit during my M.Sc. programme. With endless pleasure, I wish to express my indebtedness and deep sense of gratitude to Dr. Ravinder Kaur, Director (Acting), IARI for his support, encouragement and inspiration. My sincere thanks are to Dr. K.V. Prabhu, Joint Director (Research), IARI for providing infrastructural facilities during the period. I extend my sincere thanks to Dr. R.K. Jain, Dean and Joint Director (Education) for all the academic support and encouragement extended throughout my M.Sc., programme. I am grateful to Dr. A.K. Singh, Head, Division of Genetics for his help extended during the period of my study. I also extend my sincere thanks to Dr. Vinod, Professor, Division of Genetics, for his intellectual suggestions, constant inspiration and encouragement during the course of this investigation. I take this opportunity to sincerely thank Dr. H.S. Gupta, former Director, IARI for his valuable suggestions. I am indebted to T. Nepolean, Senior Scientist, Division of Genetics and Co- chairman of my Advisory Committee and Dr. S.K. Jha, Scientist, Division of Genetics for their generous advice. I feel immense pleasure to convey my heartfelt thanks to Dr. Supradip Saha, Senior Scientist,Division of Agricultural Chemicals, IARI, New Delhi; Dr. Amol Kumar Solanke, Senior Scientist, NRCPB, New Delhi. I humbly place on record my respect and gratitude to Dr. M. Vignesh, Scientist, Division of Genetics, for his valuable guidance and scientific suggestions which have greatly facilitated the production of this thesis. He was always there in all my need and helped his best whenever I seek for it. I am indebted to him for the critical reviewing of the manuscript. I am immensely indebted to Rajkumar sir and Aanchal mam for their constant guidance and help during the entire period of my research work. I feel privileged to express my heartier gratitude to my beloved teachers, and staff members, Ishwar sir, Lalji sir, Division of Genetics, for their immense help in successfully arranging out of this study. I wish to express my deepest sense of gratitude and indebtedness to Dr. Rajesh Kumar, Technical officer, Maize Genetics Unit, who helped me a lot in every aspect of study. I want to thank Diwakar ji and R.P. Singh ji who provided all kind of support during my data collection and field trials. It gives me immense pleasure to mention names of Rahmat sir, Rajat sir, Anil bhaiya and Ajay bhaiya, whose constant help and collective efforts have been reflected in the completion of this venture. I also extend my thanks to Jalil Ji, Mukhtyar Singh ji and Sunita ji for their help during the execution of experiments and data collection. It’s my pleasure to thank my dear seniors Mallikarjuna, Sarika, Rahul, Vidyasagar, Mukesh, Subhash, Amit, Soma and Manu for their constant help and guidance during my research work. Candid thanks to my juniors, Nitish, Premchand, Prahlad, Yankappa, Shyamsundar and Damodar, and my friends Virendra, Sunil, Arvind, Pradeep, Amarkant, Ashish, Neeraj, Gaurendra, and Ganesh for their support and help in various forms and other gestures. Diction is not enough to express my unboundful gratitude and affection to my beloved parents Shri. Madan Lal and Smt. Nabbo Devi for bringing me up in the best of ways, for rendering me the best of education, for nurturing in me the best of ideals and for helping me to see the best of times. There is no match to the love I show to my family members, Pramod, Manoj, Rohit, Keshav, Seema di, Jyoti di, Asha di, Saroj di, Usha, Archana, Poonam, Hemlata, Pooja, Monika and little flowers of our garden Vipin, Vishal, Prashant, Kannu,Abhishek, Sapna, Mangal and Rakhi who made my life the happiest throughout. I must express my gratitude to my mamaji Panchu Lal, for his continued support, unwavering love and encouragement. My vocabulary utterly fails in expressing my love to my relatives and friends for their support during all the years of study. Finally, the financial assistance provided by IARI in the form of Fellowship during the tenure is gratefully acknowledged. Though many have not been mentioned, none are forgotten. Date: 29.07.2015 Place: New Delhi (Brijesh Mehta) CONTENTS S. No. Title Page No. 1 Introduction 1 2 Review of Literature 4 3 Materials and Methods 23 4 Results 30 5 Discussion 41 6 Summary and Conclusion 54 Abstract lkjka'k Bibliography i - xx LIST OF TABLES Table After Title No. page No. 2.1 List of various endosperm mutants in maize 7 List of genotypes used for molecular characterisation using 3.1 27 SSR markers ANOVA for morphological and quality traits of sh2sh2-based 4.1 31 sweet corn hybrids (Environment-I) ANOVA for morphological and quality traits of sh2sh2-based 4.2 31 sweet corn hybrids (Environment-II) ANOVA for morphological and quality traits of sh2sh2-based 4.3 31 sweet corn hybrids (Environment-III) ANOVA for morphological and quality traits of sh2sh2/su1su1- 4.4 31 based sweet corn hybrids (Environment-I: First date of sowing) ANOVA for morphological and quality traits of sh2sh2/su1su1- 4.5 based sweet corn hybrids (Environment-II: Second date of 31 sowing) ANOVA for morphological and quality traits of sh2sh2/su1su1- 4.6 based sweet corn hybrids (Environment-III: Third date of 31 sowing) Combined ANOVA for morphological traits of sh2sh2-based 4.7 31 sweet corn hybrids Combined ANOVA for morphological traits of sh2sh2/su1su1- 4.8 31 based sweet corn hybrids Combined ANOVA for quality trait of sh2sh2-based sweet corn 4.9 31 hybrids Combined ANOVA for quality trait of sh2sh2/su1su1-based 4.10 31 sweet corn hybrids Mean performance of sh2sh2-based sweet corn hybrids for 4.11 31 morphological traits Mean performance of sh2sh2/su1su1-based sweet corn hybrids 4.12 31 for morphological traits Mean performance of sh2sh2-based sweet corn hybrids for 4.13 33 quality trait Mean performance of sh2sh2/su1su1-based sweet corn hybrids 4.14 34 for quality trait Correlation among various morphological and quality 4.15 characters of sh2sh2-based sweet corn hybrids over 37 environments Correlation among various morphological and quality 4.16 characters of sh2sh2/su1su1-based sweet corn hybrids over 37 environments Standard heterosis for cob yield and fodder yield among 4.17 37 sh2sh2-based sweet corn hybrids over environments Standard heterosis for cob yield and fodder yield among 4.18 37 sh2sh2/su1su1-based sweet corn hybrids over environments 4.19 List of SSR markers used in the study and their properties 39 Dissimilarity matrix among the 48 genotypes (1-48) used in the 4.20 39 study LIST OF FIGURES Figure After Title No. Page No. 2.1 A simplified illustration of starch biosynthesis pathway in 7 maize 4.1 Selected cobs of promising sh2sh2- and sh2sh2/su1su1- 33 based sweet corn hybrids 4.2 SSR polymorphism among sweet corn inbreds 39 4.3 Cluster analyses depicting genetic relationships among 48 39 inbreds 4.4 Principal Coordinate Analyses depicting genetic 40 relationships among 48 inbreds 5.1 Comparison of performance of promising experimental 48 hybrids over checks for A: cob yield and B: brix. 5.2 Comparison of performance of promising experimental 48 hybrids over checks for A: fodder yield and B: anthesis. ACRONYMS µg : microgram µl : microlitre bp : base pair AICMIP : All India Coordinated Maize Improvement Project ANOVA : Analysis of Variance CTAB : Cetyl Trimethyl Ammonium Bromide DAP : Days After Pollination DNA : Deoxyribo Nucleic Acid dNTP : deoxynucleotide 5’ triphosphate EDTA : Ethylene Diamine Tetra-acetic Acid EtBr : Ethidium Bromide HPLC : High Performance Liquid Chromatography IARI : Indian Agricultural Research Institute ICAR : Indian Council of Agricultural Research IIMR : Indian Institute of Maize Research MGU : Maize Genetics Unit PCR : Polymerase Chain Reaction PCoA : Principal Coordinate Analysis PIC : Polymorphism Information Content SNP : Single Nucleotide Polymorphism SSR : Simple Sequence Repeats 1 INTRODUCTION Maize (Zea mays ssp. mays L., 2n=20), considered as queen of the cereals, is one of the most important cereal crops in the world next only to wheat and rice. Globally, annual maize production is about 1016.73 million metric tonnes, of which Asia alone produces 304.31 million metric tonnes (FAOSTAT, 2013). India is the second-most important maize growing country in Asia, and is the world's sixth largest producer and the fifth-largest consumer of maize (Prasanna, 2014). During 2013-14, India produced 24.35 million metric tonnes of maize from 9.4 million hectares (www.indiastat.com). Nearly 23% of the maize produced in India, is used for human food and around 63% is utilized as poultry- and animal- feed (Yadav et al., 2014). It is a staple food crop in many tropical African and Asian countries (Shiferaw et al., 2011). Of the various specialty corns, sweet corn (Z. mays ssp. mays var. saccharata) has market potential not only in domestic market, but in international market as well. Sweet corn is a type of corn with thin layer of pericarp and is consumed at immature stages of endosperm development at ~20-22 days after fertilization. Sweet corn has emerged as one of the popular choices worldwide, both as fresh and processed vegetable. It is also an important source of fibre, minerals, and vitamins. Generally, sweet corn is marketed in three distinct forms: fresh, canning, and freezing. Fresh sweet corn products such as sweet corn milk and soups are gaining popularity in many countries. Sweet corn ears are eaten green as highly prized fresh products like baby corn. Immature kernels are ground to produce the confection pinnole and as a fermentable source for the production of an alcoholic drink - chichi. The adoption of new sweet corn based products has increased its consumption and has helped to further expand the market. Global import of frozen sweet corn was valued over US $332 million, while the same for the preserved sweet corn was estimated as US $876 million during 2010. Countries such as US, Hungary, France, New Zealand, Belgium and Thailand are the leading exporters of the sweet corn based products. In Asian countries, Japan and Taiwan occupies approximately 35,000 and 20,000 ha area under sweet corn, respectively. The demand of sweet corn is increasing tremendously in the last few years due to the increasing urbanization.
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