Phylogenetic Relationships Within Columnea Section
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PHYLOGENETIC RELATIONSHIPS WITHIN COLUMNEA SECTION ANGUSTIFLORAE : INSIGHTS INTO FORCES DRIVING SPECIATION by Lacie Janelle Schulte A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science in Biology Boise State University August 2012 © 2012 Lacie Janelle Schulte ALL RIGHTS RESERVED BOISE STATE UNIVERSITY GRADUATE COLLEGE DEFENSE COMMITTEE AND FINAL READING APPROVALS of the thesis submitted by Lacie Janelle Schulte Thesis Title: Phylogenetic Relationships within Columnea Section Angustiflorae : Insights into Forces Driving Speciation Date of Final Oral Examination: 01 June 2012 The following individuals read and discussed the thesis submitted by student Lacie Janelle Schulte, and they evaluated her presentation and response to questions during the final oral examination. They found that the student passed the final oral examination. James F. Smith, Ph.D. Chair, Supervisory Committee Steven J. Novak, Ph.D. Member, Supervisory Committee Merlin White, Ph.D. Member, Supervisory Committee John L. Clark, Ph.D. Member, Supervisory Committee The final reading approval of the thesis was granted by James F. Smith, Ph.D., Chair of the Supervisory Committee. The thesis was approved for the Graduate College by John R. Pelton, Ph.D., Dean of the Graduate College. DEDICATION To my parents, Don and Vickie, and my brother, Alex, for their unending support in all my endeavors. To Elizabeth Bader, for always encouraging me to spread my wings and fly. iv ACKNOWLEDGEMENTS This thesis would not have been possible without the contributions and support of many people, but none were more important than my major advisor, Dr. James F. Smith. I would like to thank Jim first for giving me this opportunity despite my lack of experience in both botany and phylogenetics. His passion and expertise in both fields made the learning process much easier. I am grateful that Jim has pushed me out of my comfort zone to become a better scientist and writer. I cannot express how much his patience and guidance have meant to me throughout this whole process. I would also like to thank my committee members for their assistance throughout this process: Dr. Stephen J. Novak for his insightful suggestions and explanations; Dr. Merlin White for his comments on brevity; and Dr. John L. Clark for his hours collecting plant material and knowledge of Columnea morphology. This research was made possible by the many donations of plant material. Both Dr. James F. Smith and Dr. John L. Clark have spent countless hours collecting material, and the majority of specimens sampled came from their collections. I would also like to thank Larry Skog, Nancy and Jerry Kast, Bob Stewart, Bill Price, Carol Ann Bonner, Julie Mavity-Hudson, and Karen Cichocki, who have graciously donated plant material. In addition, I would like to thank the Selby Botanical Garden, Missouri Botanical Garden, Smithsonian Institute, National Herbarium of Colombia, University of the West v Indies, and Institute of Jamaica Herbaria for donating material and allowing us to photograph their specimens. I was also given the opportunity to do field work in Jamaica. This field work would not have been possible without Keron Campbell and Judeen Meikle at the Institute of Jamaica. Keron and Judeen were gracious enough to accompany us into the field and help us find our way around Jamaica. I would also like to thank my fellow lab members. Shandra Jeffries and Maggie Ooi made this thesis possible by helping gather molecular data, georeferencing herbarium specimens, and making work fun each and every day. Thank you also to Danielle Clay who helped me with lab techniques and made me feel welcome when I first arrived at Boise State and Patrick Kolar for all his help with GIS. I would also like to say thank you to the many individuals who have made my time at Boise State a memorable experience: Morgan Peters, Emma Wilson, Jessie Sherburne, Allison Korte, and many other friends and colleagues in the graduate program. I must also acknowledge both Jane Wattrus and Gerald Cizadlo for their help in applying to the graduate program at Boise State. Without their initial encouragement and continued support, I would not have gone to graduate school. No acknowledgement section would be complete without my family and friends. I am lucky to have an amazing support group across the country that has seen me through this entire process. I could not name them all, but I must say thank you to the people who heard me complain during the hard times but do not hear thank you enough: Mom and Dad, Al, Kathy and Randy, Linda and Jeff, Elizabeth, Kelly, and Caitie - thank you so much for all your love and support. vi This research was made possible by funding from the National Science Foundation and the Gesneriad Society Elvin McDonald Research Endowment Fund. I would also like to thank the Gesneriad Society for awarding me the Student Travel Grant that allowed me to attend the 2011 Gesneriad Convention and meet so many of these Gesneriad enthusiasts who keep these plants growing and make Gesneriaceae research possible. vii ABSTRACT Determining the specific factors that played a role in speciation previously took extensive resources that made such studies nearly intractable. Despite the difficulties presented by speciation studies, we are still interested in determining what forces drive the process of evolution to gain a better understanding of divergence among species. Advances in technology allow for a new approach to speciation studies, beginning with molecular phylogenetic analyses that identify the species within a monophyletic clade and generate a species-level phylogeny. Molecular data are an independent source of data and provide a phylogeny to map both morphological characters and ecological parameters. Identifying patterns among phylogenetic studies, morphological characters, and ecological variables highlight possible forces driving speciation. Morphological characters have previously provided the foundation for phylogenetics. However, convergence among characters has made phylogenetic studies difficult based on morphology alone. Molecular phylogenetic analyses provide better insight into relationships across the family and as a result, better classification systems that are a reflection of ancestral evolutionary history rather than convergent evolution. As the largest Neotropical genus in the family Gesneriaceae, with over 200 species, Columnea has had a complex taxonomic history with classification systems based on morphological characters including vegetative, nectary, and floral characters. Most recent classification systems had classified the species of Columnea into five viii sections: sections Columnea, Collandra, Ortholoma, Pentadenia, and Stygnanthe . Section Stygnanthe encompassed eighteen species based on a similar floral morphology, characterized by small corollas that are slightly ventricose and constricted at the base, ranging in size from 1.4 to 5.2 cm in length. A well-supported topology of the species of Columnea can test the previous subgeneric classifications. The phylogenetic analyses presented here sampled 129 accessions representing 90 species within Columnea . Of the 90 species included, fifteen of the eighteen species of Stygnanthe were sampled along with an additional three species, Columnea grisebachiana , C. moorei , and C. ulei , which had not been placed in Stygnanthe but share a similar corolla morphology. Based on five chloroplast DNA (cpDNA) gene regions (trnQ-rps16 spacer, rpl32-trnL UAG spacer, rps16 intron, trnS-G spacer, and trnH-psbA spacer) and nuclear ribosomal internal transcribed spacers (ITS1 and ITS2, hereafter referred to as ITS) the results of this study indicated that the species of section Stygnanthe and the three additional species all with similar corolla morphologies, belong in three separate clades within Columnea . Three of the species from Stygnanthe (C. moesta, C. ultraviolacea , and C. xiphoidea ), including the type species and one of the additional species ( C. moorei ), were separated from the remaining twelve species of Stygnanthe that were tested. Because the type species ( C. moesta ) moved to another clade, most of the species of the former section Stygnanthe and one of the additional tested species ( C. ulei ) belong within section Angustiflorae . The other tested species, C. grisebachiana , fell out into a third separate clade. The results indicate that within Columnea small corollas that are slightly ventricose and constricted at the base are the result of convergent evolution. ix Molecular phylogenetic analyses with five cpDNA gene regions and ITS provided phylogenetic support for seven monophyletic clades within Columnea but failed to resolve species-level relationships within clades. Additional molecular phylogenetic analyses were conducted to resolve species level relationships with 36 accessions, representing thirteen of fifteen species within Angustiflorae . The external transcribed spacer (ETS) was added to the sequences from the five cpDNA gene regions and ITS from the genus wide analysis to generate a well-resolved species level phylogeny. In addition, two low-copy nuclear gene regions, glyceraldehyde 3-phosphate dehydrogenase (G3pdh ) and NADP-dependent isocitrate dehydrogenase ( idh ) were included to boost phylogenetic support of the major branching events within the section. This study used these molecular phylogenetic analyses along with morphological characters and climatic variables to determine driving forces of