University of South Dakota USD RED Honors Thesis Theses, Dissertations, and Student Projects 2017 The Old, the Young, and the Dehydrated: comparing dehydration tolerance of ‘Gransden’ and ‘Villersexel’ ecotypes of Physcomitrella patens Katherine Bollinger University of South Dakota Follow this and additional works at: https://red.library.usd.edu/honors-thesis Recommended Citation Bollinger, Katherine, "The Old, the Young, and the Dehydrated: comparing dehydration tolerance of ‘Gransden’ and ‘Villersexel’ ecotypes of Physcomitrella patens" (2017). Honors Thesis. 1. https://red.library.usd.edu/honors-thesis/1 This Honors Thesis is brought to you for free and open access by the Theses, Dissertations, and Student Projects at USD RED. It has been accepted for inclusion in Honors Thesis by an authorized administrator of USD RED. For more information, please contact [email protected]. THE OLD, THE YOUNG, AND THE DEHYDRATED: COMPARING DEHYDRATION TOLERANCE OF ‘GRANSDEN’ AND ‘VILLERSEXEL’ ECOTYPES OF PHYSCOMITRELLA PATENS by Katherine Bollinger A Thesis Submitted in Partial Fulfillment Of the Requirements for the University Honors Program ___________________________________________________________ Department of Biology The University of South Dakota December 2017 The members of the Honors Thesis Committee appointed to examine the thesis of Katherine Bollinger find it satisfactory and recommend that it be accepted. _________________________________________________ Karen L. Koster, Ph.D. Professor of Biology Director of the Committee _________________________________________________ Bernard W. M. Wone, Ph.D. Assistant Professor of Biology _________________________________________________ Kumudu N. Rathnayake Ph.D. Candidate in Biology ABSTRACT The Old, the Young, and the Dehydrated: comparing dehydration tolerance of ‘Gransden’ and ‘Villersexel’ ecotypes of Physcomitrella patens Katherine Bollinger Director: Dr. Karen L. Koster, Ph, D. Plant tolerance of environmental stresses such as water deficit depends on genetic factors that can vary in response to the climate in which the plants have evolved. Such genetically distinct populations of a species that live in different regions are referred to as ecotypes. The purpose of this research was to test the dehydration tolerance of two ecotypes of the moss species Physcomitrella patens, one from Little Gransden, England, which has been used for decades for research, and another more recently cultured ecotype from Villersexel, France. Little Gransden receives nearly half the yearly precipitation of Villersexel, which could have led to the ‘Gransden’ ecotype becoming more dehydration tolerant than the ‘Villersexel’ ecotype. To test my hypothesis, I allowed samples containing contained both protonemata and gametophores of each ecotype to equilibrate to a range of relative humidities (RH), measured the extent of dehydration they experienced, and then assessed their ability to survive dehydration using chlorophyll fluorescence as a measure of metabolic recovery after rehydration. The data indicate that the ‘Gransden’ ecotype survived and recovered from equilibration to RH as low as 89% RH, while the ‘Villersexel’ ecotype was less tolerant of dehydration, only surviving equilibration down to 95% RH. Keywords: Physcomitrella patens, dehydration tolerance, chlorophyll fluorescence, ecotypic variation, ‘Gransden’, ‘Villersexel’ TABLE OF CONTENTS LIST OF FIGURES ................................................................................................................................... vi LIST OF TABLES .................................................................................................................................... vii ACKNOWLEDGEMENTS ................................................................................................................... viii INTRODUCTION ...................................................................................................................................... 1 MATERIALS AND METHODS ........................................................................................................... 13 RESULTS .................................................................................................................................................. 21 DISCUSSION AND CONCLUSION .................................................................................................... 35 APPENDIX ............................................................................................................................................... 42 BIBLIOGRAPHY .................................................................................................................................... 50 v LIST OF FIGURES Figure 1: The life cycle of P. patens ................................................................................................. 4 Figure 2: Origin of ‘Gransden’ and ‘Villersexel’ moss ecotypes ........................................ 10 Figure 3: Rate of water loss of VX and GR samples over saturated KNO3 ................... 22 Figure 4: Rate of water loss of VX and GR samples over saturated MgSO4 .................. 22 Figure 5: Rate of water loss of VX and GR samples over saturated KCl ........................ 23 Figure 6: Rate of water loss of VX and GR samples over saturated NaCl ...................... 23 Figure 7: Rate of water loss of VX and GR samples over saturated MgCl2 ................... 24 Figure 8: Average final water content (gH2O/gDW) for VX and GR at five RH levels after 144 h equilibration period .................................................................................................... 26 Figure 9: Adjusted average final water content (gH2O/gDW) for VX and GR at five RH levels after 144 h equilibration period ................................................................................ 26 Figure 10: The mean chlorophyll fluorescence (Fv/Fm) measurements during rehydration for the ‘Villersexel’ ecotype .................................................................................... 31 Figure 11: The mean chlorophyll fluorescence (Fv/Fm) measurements during rehydration for the ‘Gransden’ ecotype ...................................................................................... 31 Figure 12: Survival of VX and GR ecotypes after equilibration to a range of RH ....... 33 Figure A1: Chlorophyll fluorescence (Fv/Fm) measurements during rehydration for Trial 1 of the ‘Villersexel’ ecotype ................................................................................................. 44 Figure A2: Chlorophyll fluorescence (Fv/Fm) measurements during rehydration for Trial 1 of the ‘Gransden’ ecotype ................................................................................................... 44 Figure A3: Chlorophyll fluorescence (Fv/Fm) measurements during rehydration for Trial 2 of the ‘Villersexel’ ecotype ................................................................................................ 46 Figure A4: Chlorophyll fluorescence (Fv/Fm) measurements during rehydration for Trial 2 of the ‘Gransden’ ecotype ................................................................................................... 46 Figure A5: Chlorophyll fluorescence (Fv/Fm) measurements during rehydration for Trial 3 of the ‘Villersexel’ ecotype ................................................................................................. 48 Figure A6: Chlorophyll fluorescence (Fv/Fm) measurements during rehydration for Trial 3 of the ‘Gransden’ ecotype ................................................................................................... 48 vi LIST OF TABLES Table 1: Climatological data of Villersexel, France and Little Gransden, England ....... 8 Table 2: Control chlorophyll fluorescence (Fv/Fm) measurements of healthy ‘Gransden’ ecotype tissue prior to dehydration ...................................................................... 29 Table 3: Control chlorophyll fluorescence (Fv/Fm) measurements of healthy ‘Villersexel’ ecotype tissue prior to dehydration .................................................................... 29 vii ACKNOWLEDGEMENTS This study was funded through a Council for Undergraduate Research and Creative Scholarship (CURCS) Research Grant, provided by the University of South Dakota. I would like to thank the council for awarding me with the opportunity and means to complete this research project. I would also like to thank my husband, family, and friends for the continuous encouragement throughout the process of planning, preparing, researching, writing, and editing of my thesis. It is because of their constant support that I was able to persevere through the challenging times. Next, I would like to thank Clay Lippert for researching alongside me in the Koster laboratory. I enjoyed the time we spent together as colleagues and appreciated learning alongside you. Lastly, I would like to thank Dr. Koster for her constant guidance, assistance, and direction. It is because of Dr. Koster’s passion that I initially became interested in research and her dedication to teaching that I learned about plant physiology and improved my scientific writing. She spent countless hours assisting me in the lab, explaining plant physiology, challenging my writing, and helping with revisions of my thesis. I cannot thank her enough. viii INTRODUCTION Life as we know it would not be possible without plants. Plants provide many items we utilize in our daily life such as pharmaceuticals, food, textiles,