Brigham Young University BYU ScholarsArchive All Theses and Dissertations 2016-05-01 The aP th to Understanding Salt Tolerance: Global Profiling of Genes Using Transcriptomics of the Halophyte Suaeda fruticosa Joann Diray Arce Brigham Young University Follow this and additional works at: https://scholarsarchive.byu.edu/etd Part of the Microbiology Commons BYU ScholarsArchive Citation Arce, Joann Diray, "The aP th to Understanding Salt Tolerance: Global Profiling of Genes Using Transcriptomics of the Halophyte Suaeda fruticosa" (2016). All Theses and Dissertations. 6355. https://scholarsarchive.byu.edu/etd/6355 This Dissertation is brought to you for free and open access by BYU ScholarsArchive. It has been accepted for inclusion in All Theses and Dissertations by an authorized administrator of BYU ScholarsArchive. For more information, please contact [email protected], [email protected]. The Path to Understanding Salt Tolerance: Global Profiling of Genes Using Transcriptomics of the Halophyte Suaeda fruticosa Joann Diray Arce A dissertation submitted to the faculty of Brigham Young University in partial fulfillment of the requirements for the degree of Doctor of Philosophy Brent L. Nielsen, Chair Mark J. Clement R. Paul Evans Joel S. Griffitts Peter J. Maughan Department of Microbiology and Molecular Biology Brigham Young University May 2016 Copyright © 2016 Joann Diray Arce All Rights Reserved ABSTRACT The Path to Understanding Salt Tolerance: Global Profiling of Genes Using Transcriptomics of the Halophyte Suaeda fruticosa Joann Diray Arce Department of Microbiology and Molecular Biology, BYU Doctor of Philosophy Salinity is a major abiotic stress in plants that causes significant reductions in crop yield. The need for improvement of food production has driven research to understand factors underlying plant responses to salt and mechanisms of salt tolerance. The aim of improving tolerance in traditional crops has been initiated but most crops can only tolerate a limited amount of salt in their systems to survive and produce biomass. Studies of naturally occurring high salt-tolerant plants (halophytes) are now being promoted for economic interests such as food, fodder or ecological reasons. Suaeda fruticosa, a member of the family Chenopodiaceae, belongs to a potential model halophyte genus for studying salt tolerance. However, published reports on the identification of genes, expression patterns and mechanisms of salinity tolerance in succulent halophytes are very limited. Next generation RNA-sequencing techniques are now available to help characterize genes involved in salinity response, along with expression patterns and functions of responsive genes. In this study, we have optimized the assembly of the transcriptome of S. fruticosa. We have annotated the genes based on their gene ontology characteristics and analyzed differential expression to identify genes that are up- and down- regulated in the presence of salt and have grouped the genes based on their putative functions. We also have provided evidence for groups of transcription factors that are involved in salt tolerance of this species and have identified those that may affect the regulation of salt tolerance. This work elucidates the characterization of genes involved in salinity tolerance to increase our understanding of the regulation of salt in a succulent halophyte. Keywords: Suaeda fruticosa, halophytes, salt tolerance, transcriptome, RNA-seq, salinity ACKNOWLEDGEMENTS Words cannot fully express my appreciation to my Heavenly Father for all the talents, guidance and blessings He has given me and my family as I go through this graduate experience. I would like to express my gratitude and forever appreciation to my mentor, Dr. Brent L. Nielsen for his full support, words of wisdom and exemplary dedication to my graduate experience. He has helped me succeed despite of my failures, limitations and challenges. He has given me an outstanding example of humility, hard work and faith that everything will work out in the end. I would like to thank my excellent committee members, Dr. Griffitts, Dr. Maughan, Dr. Evans, Dr. Prince and Dr. Clement for their invaluable critiques and for allowing me to grow and become a better scientist. I would like to specially thank Dr. Mark J. Clement for his expertise and encouragement to learn Bioinformatics. I would like to thank my BYU colleagues and collaborators that made significant contributions to my projects: Drs. Ajmal Khan and Bilquees Gul for the halophyte project, Bin Liu for the Twinkle project, Huan Kang for the proteomics analysis, Justin Page, Paul Bodily and Stanley Fujimoto for their bioinformatics expertise, Anton Suvorov for his evolutionary biology expertise, Collin Hansen, Stewart Morley and all Nielsen lab undergraduates for their help in the lab. I would also want to express my gratitude to my parents, Jorge and Reneca Diray for teaching me that learning does not stop, and to my siblings and in-laws for their support and encouragement despite being far away from them. Finally, I would like to dedicate this dissertation to the love of my life: my husband Carlo and my children born in-between writing papers and grants, Travis and Arabella. TABLE OF CONTENTS TITLE PAGE ................................................................................................................................... i ABSTRACT .................................................................................................................................... ii ACKNOWLEDGEMENTS ........................................................................................................... iii TABLE OF CONTENTS ............................................................................................................... iv LIST OF TABLES ......................................................................................................................... ix LIST OF FIGURES ........................................................................................................................ x CHAPTER 1: Halophyte Transcriptomics: Understanding Mechanisms of Salinity Tolerance .... 1 ABSTRACT ............................................................................................................................................. 2 INTRODUCTION .................................................................................................................................... 2 Transcriptome Sequencing Overview .................................................................................................. 3 Applications of RNA Studies ............................................................................................................... 4 NGS Approaches for Salt-Tolerance Studies ....................................................................................... 7 Genes Involved in General Metabolism ............................................................................................... 9 Genes for Cell Maintenance ................................................................................................................. 9 Genes Encoding Plant Hormones ....................................................................................................... 13 Genes Encoding Ion Transporters ...................................................................................................... 14 Regulatory Molecules ........................................................................................................................ 16 LEA Protein Coding Genes ................................................................................................................ 17 Other Genomic Elements ................................................................................................................... 17 Pathways ............................................................................................................................................ 18 CONCLUSIONS AND FUTURE DIRECTIONS ................................................................................. 19 BIBLIOGRAPHY .................................................................................................................................. 20 iv CHAPTER 2: Suaeda fruticosa, a Potential Model Halophyte for Salt Tolerance Research ....... 26 REFERENCES ....................................................................................................................................... 31 CHAPTER 3: Optimization of de novo Transcriptome Assembly of the Halophyte Suaeda fruticosa Using Clustering Methods ............................................................................................. 34 ABSTRACT ........................................................................................................................................... 35 BACKGROUND .................................................................................................................................... 36 RESULTS AND DISCUSSIONS .......................................................................................................... 39 Sequencing Method and Quality Assessment of the Reads ............................................................... 39 Normalizing Reads by k-mer Coverage ............................................................................................. 39 De novo Transcriptome Assembly ..................................................................................................... 40