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Analysis of a Wound-Induced Gene Family in Glycine Max

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Analysis of a Wound-Induced Gene Family in Glycine Max Scholars' Mine Masters Theses Student Theses and Dissertations Fall 2012 Analysis of a wound-induced gene family in Glycine max Gena Robertson Follow this and additional works at: https://scholarsmine.mst.edu/masters_theses Part of the Biology Commons, and the Environmental Sciences Commons Department: Recommended Citation Robertson, Gena, "Analysis of a wound-induced gene family in Glycine max" (2012). Masters Theses. 6940. https://scholarsmine.mst.edu/masters_theses/6940 This thesis is brought to you by Scholars' Mine, a service of the Missouri S&T Library and Learning Resources. This work is protected by U. S. Copyright Law. Unauthorized use including reproduction for redistribution requires the permission of the copyright holder. For more information, please contact [email protected]. ANALYSIS OF A WOUND-INDUCED GENE FAMILY IN GLYCINE MAX by GENA ROBERTSON A THESIS Presented to the Faculty of the Graduate School of the MISSOURI UNIVERSITY OF SCIENCE AND TECHNOLOGY In Partial Fulfillment of the Requirements for the Degree MASTER OF SCIENCE IN APPLIED AND ENVIRONMENTAL BIOLOGY 2012 Approved by Ronald L. Frank, Advisor Melanie Mormile David J. Westenberg iii ABSTRACT Gene families in plants are important in understanding genome evolution indicating when and where genome duplications and segmental duplications occurred as well as subsequent divergence and subfunctionalization. A gene family in Glycine max that encodes a WI12 protein, wound-induced protein, was found to consist of ten genes on five chromosomes. Wound-induced proteins are activated in response to wounding in plants, and the WI12 protein in particular is thought to be involved in cell wall modifications at the wound site. A variety of bioinformatics tools have been used to analyze the expansion of this family in soybean as well as identify potential functional domains in the protein. iv ACKNOWLEDGMENTS I would like to thank, first and foremost, my advisor Ronald Frank, whose support and knowledge has been invaluable. Without his help, this thesis would not have been possible. I simply could not have asked for a better advisor. I would like to thank my graduate committee members, Melanie Mormile and David Westenberg. I would like to acknowledge the Biological Sciences Department at Missouri University of Science and Technology for graduate student support. Finally, I would like to thank Ciarán Ryan-Anderson for his encouragement and formatting help. v TABLE OF CONTENTS Page ABSTRACT ...................................................................................................................... iii ACKNOWLEDGMENTS ................................................................................................ iv LIST OF ILLUSTRATIONS .......................................................................................... viii LIST OF TABLES ............................................................................................................ ix SECTION 1. INTRODUCTION .............................................................................................. 1 1.1. GLYCINE MAX .................................................................................. 1 1.2. GENE DUPLICATION AND GENE FAMILIES .............................. 3 1.3. WOUND-INDUCED PROTEIN ......................................................... 4 1.4. PROTEIN STRUCTURE AND FOLDING ........................................ 5 1.5. MEDICAGO TRUNCATULA ............................................................ 6 1.6. SELAGINELLA MOELLENDORFFII ............................................... 6 1.7. DATABASES AND TOOLS .............................................................. 6 1.7.1. Protein Databases .................................................................. 6 1.7.2. BLAST .................................................................................. 7 1.7.3. Expressed Sequence Tags ..................................................... 8 1.7.4. Phytozome............................................................................. 8 1.7.5. Sequence Alignments............................................................ 9 1.7.6. MEME……………………………………………………..10 1.7.7. Augustus ............................................................................. 11 vi 1.7.8. CELLO…………………………………………………….11 1.7.9. PLACE…………………………………………………….11 2. MATERIALS AND METHODS……………………………………………...13 2.1. IDENTIFICATION OF GENE FAMILY MEMBERS……………..13 2.2. GENE STRUCTURE PREDICTION AND EST EXPRESSION ANALYSIS…………………………………………………………13 2.3. EVOLUTIONARY ANALYSIS……………………………………14 2.4. FUNCTIONAL ANALYSIS………………………………………..16 3. RESULTS…………………………………………………………………….. 18 3.1. IDENTIFICATION OF GENE FAMILY MEMBERS……………..18 3.2. GENE STRUCTURE PREDICTION AND EST EXPRESSION ANALYSIS………………………………………………………… 19 3.3. EVOLUTIONARY ANALYSIS……………………………………24 3.4. FUNCTIONAL ANALYSIS………………………………………..27 4. DISCUSSION………………………………………………………………… 31 4.1. IDENTIFICATION OF GENE FAMILY MEMBERS……………..31 4.2. GENE STRUCTURE PREDICTION AND EST EXPRESSION ANALYSIS…………………………………………………………31 4.3. EVOLUTIONARY ANALYSIS……………………………………32 4.4. FUNCTIONAL ANALYSIS………………………………………..34 APPENDICES A. NUCLEOTIDE SEQUENCES…............................................................................. 36 B. ESTS…………………………………………………………………………….. 47 C. PHYLOGENETIC TREE FILE……………………………...…………………..60 D. PGENTHREADER AND PDOMTHREADER PREDICTIONS.…………..…..62 vii E. PLACE DATA….................................................................................................. 75 F. CELLO DATA………………………………………………………………..... 95 BIBLIOGRAPHY……………………………………………………………………... 101 VITA……………………………………………………………………………………106 viii LIST OF ILLUSTRATIONS Figure Page 3.1. Chromosome Map…………………………………………………………………. 20 3.2. ClustalW2 Protein Alignment……………………………………………………... 24 3.3. Phylogeny………………………………………………………………………..... 25 3.4. Neighbor Analysis……………………………………………………………….... 26 3.5. pGenTHREADER………………………………………………………………..... 27 3.6. pDomTHREADER……………………………………………………………..…. 28 3.7. GmWI-1 PLACE……………………………………………………………...……28 3.8. GmWI-2 PLACE……...……………………………………………………...…… 29 3.9. MEME Motifs………………………………………………………………………30 ix LIST OF TABLES Table Page 1.1. Glycine max Classification……………………………………………………......... 1 3.1. Glycine max Gene Table………………………………………………………....... 18 3.2. Medicago truncatula Gene Table………………………………………………..... 18 3.3. Selaginella moellendorffii Gene Table…………………………………………..... 19 3.4. EST Cultivars……………………………………………………………………… 21 3.5. EST Tissues……………………………………………………………………...... 22 3.6. EST Treatments…………………………………………………………………… 23 3.7. SNAP Results……………………………………………………………………... 27 1. INTRODUCTION 1.1. GLYCINE MAX Glycine max is commonly known as cultivated soybean. It is a dicotyledon crop plant found in the legume family. The classification of G. max can be seen in Table 1.1 below. Table 1.1. Glycine max Classification [1] Kingdom Plantae Plants Subkingdom Tracheobionta Vascular plants Superdivision Spermatophyta Seed plants Division Magnoliophyta Flowering plants Class Magnoliopsida Dicotyledons Subclass Rosidae Rosids Order Fabales Family Fabaceae Legumes Genus Glycine Willd. Soybean Species Glycine max Soybean G. max is one of the world’s principle food crops. Cultivation of the domesticated soybean dates back as early as 3100 years ago in East Asia [2]. It is native to North and Central China. Soybean was introduced into the United States in 1765 [3], and into Canada in 1893 [4]. Soybean is the most valuable legume crop. It is used for its seed protein and oil content, and is the main source of biodiesel. It also fixes nitrogen in the soil by way of a symbiotic relationship with the bacterium Bradyrhizobium japonicum. G. max makes up more than 55% of all oilseed production, and 80% of consumable fats and oils in the 2 United States [5]. In 2011, 3,056,032,000 bushels of soybean were produced on 73,636,000 acres in the U.S [6]. The U.S. is the main producer of soybean, making up roughly 36% of the world’s production, followed by Brazil, Argentina, China, and India, all of which comprise approximately 95% of total production [7]. The soybean genome was sequenced in 2008 through large-scale shotgun sequencing, initiated by the Department of Energy-Joint Genome Institute (DOE-JGI) Community Sequencing Program. Approximately 975 megabases (Mb) are present in 20 chromosomes, with a small additional amount found in mostly repetitive unmapped scaffolds. An initial screen of the genome resulted in the prediction of 46,430 high- confidence genes. Approximately 78% of these genes are found in the chromosome ends. Of the predicted genes, there are 31,264 that have been approximated into 12,253 gene families, and 15,166 that exist as singletons [8]. It should be noted that the number of genes families in soybean is only an estimate. Papilionoideae (the “papilionoids”) is the largest of the three subfamilies of Fabaceae (the legumes) with 476 genera and 13,860 species. Papilionoideae includes the grain legumes (beans, lentils, peanuts, etc.) [9]. A sub-clade of the papilionoids, the phaseoloids, contains Glycine, Phaseolus (bean), Cajanus (pigeon pea), and Vigna (lentil, mung bean, cowpea) [10]. Another sub-clade of the papilionoids, the galegoids (Medicago, Pisum) diverged from the phaseoloids approximately 50 million years ago (Mya) [10]. The papilionoid origin has been predicted to have occurred approximately 59 Mya by way of matK and rbcL loci analysis. The origin of legumes has been estimated to 56 Mya using fossils [11]. The Glycine-specific genome duplication has been
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