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The Regulation of Ontogenetic Diversity in Papaveraceae Compound Leaf Development

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The Regulation of Ontogenetic Diversity in Papaveraceae Compound Leaf Development The Regulation of Ontogenetic Diversity in Papaveraceae Compound Leaf Development A thesis presented to the faculty of the College of Arts and Sciences of Ohio University In partial fulfillment of the requirements for the degree Master of Science Alastair R. Plant August 2013 © 2013 Alastair R. Plant. All Rights Reserved. 2 This thesis titled The Regulation of Ontogenetic Diversity in Papaveraceae Compound Leaf Development by ALASTAIR R PLANT has been approved for the Department of Environmental and Plant Biology and the College of Arts and Sciences by Stefan Gleissberg Assistant Professor of Environmental and Plant Biology Robert Frank Dean, College of Arts and Sciences 3 ABSTRACT PLANT, ALASTAIR R., M.S., August 2013, Plant Biology The Regulation of Ontogenetic Diversity in Papaveraceae Compound Leaf Development Director of Thesis: Stefan Gleissberg The leaf is almost ubiquitous throughout land plants but due to its complex and flexible developmental program is highly morphologically variable between taxa. Description of the functions of regulatory genes key to leaf development in different evolutionary lineages allows the study of changes in developmental mechanisms through evolutionary time as a means for anatomical and morphological diversification. The roles of homologs of CINCINNATA-like TCP family genes, ARP genes, and Class I KNOX genes were investigated in two members of the Papaveraceae, a basal eudicot lineage positioned in between major angiosperm groups, by phylogenetic analysis, in situ hybridization, expression profiling by quantitative polymerase chain reaction, and virus- induced gene silencing in Eschscholzia californica and Cysticapnos vesicaria. Expression data were similar to those for homologous genes in core eudicot species, however, some gene functions found in core eudicots were not associated with basal eudicot homologs, and so have either been gained or lost from the ancestral state. This reflects the dynamism of the leaf developmental plan and its diversification through evolution. 4 DEDICATION This thesis is dedicated to curiosity. 5 ACKNOWLEDGMENTS I would like to thank those who have contributed to this body of research: Anandi Bhattacharya cloned the TCP domain of Eschscholzia californica CINCINNATA (EcCIN), Stefan Gleissberg cloned EcPHAN and performed the EcPHAN expression profile RT-PCR, and Andrea Scholz produced the Eschscholzia californica PHANTASTICA (EcPHAN) VIGS construct. I would also like to thank my advisor Stefan Gleissberg and my committee members Harvey Ballard, and Sarah Wyatt for their assistance and guidance, Oriane Hidalgo and Conny Bartholmes for training and advice, and undergraduate researchers Celeste Taylor, Avery Tucker, Brooke Johnson, Ben Imbus, Chi Zhang, Emily Usher, Jennifer Leetch, and Abby Pugel for their help in data collection and in being a source of new ideas and inspiration. This research was funded in part by an Ohio University Research Committee Grant. 6 TABLE OF CONTENTS Page Abstract ............................................................................................................................... 3 Dedication ........................................................................................................................... 4 Acknowledgments............................................................................................................... 5 List of Tables ...................................................................................................................... 8 List of Figures ..................................................................................................................... 9 Chapter 1: Introduction ..................................................................................................... 12 References ..................................................................................................................... 20 Chapter 2: Regulation of dissected leaf architecture in Eschscholzia californica By a CIN-TCP gene .................................................................................................................. 28 Abstract ......................................................................................................................... 28 Introduction ................................................................................................................... 29 Methods ........................................................................................................................ 33 Results ........................................................................................................................... 44 Discussion ..................................................................................................................... 56 References ..................................................................................................................... 59 Chapter 3: Eschscholzia californica PHANTASTICA (EcPHAN) regulates petal morphogenesis in the California Poppy ............................................................................ 65 Abstract ......................................................................................................................... 65 Introduction ................................................................................................................... 65 Methods ........................................................................................................................ 69 Results ........................................................................................................................... 71 Discussion ..................................................................................................................... 77 References ..................................................................................................................... 80 Chapter 4: Duplicated STM-like KNOX I genes act in floral meristem activity in Eschscholzia californica (Papaveraceae) .......................................................................... 86 Authors .......................................................................................................................... 86 Abstract ......................................................................................................................... 86 Introduction ................................................................................................................... 87 7 Materials and Methods .................................................................................................. 91 Results ........................................................................................................................... 95 Discussion ................................................................................................................... 110 Acknowledgements ..................................................................................................... 117 Author Contributions .................................................................................................. 117 References ................................................................................................................... 118 Supplementary Data ................................................................................................ 128 Chapter 5: Laser microdissection of Eschscholzia californica leaf primordia for comparison of gene expression between developmental stages ..................................... 134 Abstract ....................................................................................................................... 134 Introduction ................................................................................................................. 134 Method ........................................................................................................................ 138 Results ......................................................................................................................... 141 Discussion ................................................................................................................... 143 Acknowledgements ..................................................................................................... 146 References ................................................................................................................... 146 Chapter 6: Discussion ..................................................................................................... 149 References ................................................................................................................... 151 Appendix I: Cloning of putative microRNA319 homologs from Eschscholzia californica ......................................................................................................................................... 153 References ................................................................................................................... 156 Appendix II: Cloning of gibberellic acid and cytokinin metabolic genes from Eschscholzia californica ................................................................................................. 158 References ................................................................................................................... 162 8 LIST OF TABLES Page Table 1: Primers used for cloning, in situ hybridization probe preparation and quantitative PCR for CIN-TCP
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