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Natural Trait Variation for Taxonomic Classification and Breeding Potential Assessment in the Genus Camelina

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Natural Trait Variation for Taxonomic Classification and Breeding Potential Assessment in the Genus Camelina Natural trait variation for taxonomic classification and breeding potential assessment in the genus Camelina by Jerry Wu A thesis submitted to the Faculty of Graduate and Postdoctoral Affairs in partial fulfillment of the requirements for the degree of Master of Science in Biology Carleton University Ottawa, Ontario © 2016, Jerry Wu Abstract Camelina sativa (L.) Crantz or camelina has been the subject of renewed interest as a novel alternative oilseed crop suitable for uses in biofuel, food, and industrial chemical applications as well as sustainable agricultural practices. Camelina’s development as an oilseed crop is currently limited by a short breeding history and a complicated allohexaploid genome structure, which hinders traditional breeding and genetic modification approaches. Therefore, our study takes an alternative approach, examining natural (phenotypic) variation in an assortment of quantitative traits of 23 accessions representing camelina and four of its crop wild relatives (CWRs) to further our understanding of the taxonomic classification and breeding potential within the Camelina genus. For our first objective, to determine key traits for discriminating taxa in the genus, principal components analysis (PCA) and linear discriminants analysis (LDA) were conducted to understand the patterns of phenotypic variation observed. It was determined that C20:1 and C22:1 fatty acid traits were the most informative variables in discriminating eight group membership identifications of the Camelina genus in this study. Permutational MANOVAs and Welch’s t-tests conducted across the eight group membership identifications as well as 23 accessions for the quantitative traits were consistent with the clustering analyses from PCA and LDA. Also, our investigations into varying morphological traits within the genus combined with patterns of variation observed for seed oil characteristics suggest that C. hispida Boiss. is a potential parental ancestor of camelina. For our second objective, we established three criteria measurements to screen for candidate plant lines with desirable fatty acid profiles for further development in breeding programmes for biofuel, food, and industrial chemical applications. We identified individual ii lines each of C. sativa (ID# S-239), C. hispida (ID# HG-240), and diploid C. microcarpa Andrz. ex DC. (ID# M2-246) as well as five C. rumelica Velen. accessions for development in the aforementioned applications, respectively. This study provides the foundation for subsequent research on basic plant biology and directions for breeding programmes in an allopolyploid oilseed crop. iii Acknowledgements I would like to first thank my main supervisor, Dr. Owen Rowland, for not only providing me with the opportunity to carry out a novel and interesting research project but also for his award-winning mentorship. If there’s one thing to take away from the invaluable experiences obtained throughout this degree, it was your advice to be persistent and I am grateful I took it not only to see this project through but as an essential skill for a successful career in science. I would next like to thank my co-supervisor, Dr. Sara Martin, for providing us with all the necessary resources required at ORDC and her contributions in running flow cytometry to generate the large dataset in this study. Thanks to Dr. Tyler Smith for helping me learn RStudio and accompanying statistical analyses to satisfy my curiosity in using a programming language to efficiently process and organize my dataset. I would also like to thank my M.Sc. Committee Members, Dr. Shelley Hepworth and Dr. Douglas Johnson, for their support and guidance. Thanks to Dr. Farah Hosseinian and her team for allowing me to use their N2 analyzer for protein content measurements. A special thank-you to members of both the Rowland and Martin Labs; Dr. Ian Pulsifer for his help with gas chromatography, Sarah Endenburg for her help with microscopic morphological measurements, Tracey James, Rylee Oosterhuis, and Connie Sauder for assistance with plant growth and pollination, Dr. Jhadeswar Murmu for general lab skills, and everyone else for their help and good spirits. I would also like to thank the administrative staff at Carleton University and greenhouse staff at ORDC for making the completion of this M.Sc. degree a lot smoother. Last, but certainly not least, I dedicate this achievement to my family for their positive and unwavering support. Jei, Ryan, Jiash, Lisa, Madz, and Joce: on top of all your help and iv useful advice, thank you for the constant reminder that there is life outside of research. Mom and Dad: thank you for genuine unconditional love through thick and thin and providing me with endless opportunities, of which I shamefully admit, sometimes take for granted. This simply could not have been done without love and support from all of you. v Statement of Contribution I, Jerry Wu, performed all the experiments and generated all the material reported in this thesis with the exception of the following: 1. Sowing and vernalization of plant accessions was done by Tracey James (Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa). 2. Flower and pod morphological measurements via microscopy was done by Sarah Endenburg (Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa). 3. Dr. Sara Martin (Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa) contributed in measuring DNA content of plant accessions via flow cytometry. 4. LDA biplot functions were written in R by Dr. Tyler Smith (Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa). vi Table of Contents Abstract ........................................................................................................................................... ii Acknowledgements ........................................................................................................................ iv Statement of Contribution .............................................................................................................. vi Table of Contents .......................................................................................................................... vii List of Figures ................................................................................................................................ ix List of Tables .................................................................................................................................. x List of Appendices ......................................................................................................................... xi List of Abbreviations ................................................................................................................... xiii 1.0 Introduction & Literature Review ........................................................................................ 1 1.1 Incentives for more sustainable agricultural practices ..................................................... 1 1.2 Camelina as an alternative oilseed crop ........................................................................... 2 1.3 Camelina for industrial applications ................................................................................ 3 1.4 Camelina and genetic tools for crop development ........................................................... 7 1.5 Implications of natural variation in crop research ............................................................ 8 1.6 Natural variation in camelina ......................................................................................... 10 1.7 Current advances in camelina genome structure ............................................................ 11 1.8 The Camelina genus ....................................................................................................... 12 1.9 Thesis objective statements ............................................................................................ 13 2.0 Methods.............................................................................................................................. 16 2.1 Plant accessions and growth conditions ......................................................................... 16 2.2 Nuclear DNA content analysis by flow cytometry (FCM) ............................................ 19 2.3 Agronomic trait measurements ...................................................................................... 20 2.4 Pod and flower measurements........................................................................................ 21 2.5 Trichome measurements ................................................................................................ 22 2.6 Seed weight measurements ............................................................................................ 23 2.7 Protein content measurements ........................................................................................ 23 2.8 Seed oil trait measurements............................................................................................ 23 2.9 Fatty acid calculations .................................................................................................... 26 2.10 Data processing and statistical analysis ......................................................................... 27 vii 3.0 Results ...............................................................................................................................
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