Characterization of Cold and Short Day Acclimation in Grape Genotypes of Contrasting Freezing Tolerance DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Trudi Nadia Lavinia Grant Graduate Program in Horticulture and Crop Science The Ohio State University 2012 Dissertation Committee: Imed Dami, Advisor Rebecca Lamb Jim Metzger Ray Miller Eric Stockinger Copyrighted by Trudi Nadia Lavinia Grant 2012 Abstract Grapes are temperate crops, frequently damaged by freezing temperatures. The economic losses that result from freezing injury are major problems for grape and wine industries in cold regions. Our central hypothesis is that soluble sugars in grape tissues provides increased freezing tolerance (FT) and the larger sugar, raffinose, accumulates earlier and to higher amounts in cold-tolerant than in cold-sensitive cultivars in response to cold temperature and short-day (SD) photoperiod. The central objective of this dissertation was to characterize the morphological, physiological and biochemical changes in response to cold acclimation (CA) and SD photoperiod. The specific objectives were to: 1) characterize the morphological, physiological and biochemical changes induced by low temperature in cold-tolerant and cold-sensitive Vitis species under controlled conditions; 2) characterize the morphological, physiological and biochemical changes induced by SD photoperiod in cold-tolerant and cold-sensitive Vitis species under controlled conditions and 3) characterize the morphological, physiological and biochemical changes induced by low temperature and SD photoperiod in cold- tolerant and cold-sensitive Vitis species under field conditions. In response to temperature under controlled environs, grape (Vitis spp) cultivars ‘Frontenac’, ‘Couderc 3309’, ‘Concord’, ‘Cabernet Franc’, ‘Traminette’ and ‘Seyval’ were evaluated. Shoot growth slowed under cold temperature regimes in all cultivars except ‘Concord’. Under the non-acclimating temperature regime, raffinose ii concentrations were low and similar among cultivars, whereas under CA temperature regimes raffinose accumulation was generally higher and cold-tolerant cultivars accumulated higher concentrations than did cold-sensitive cultivars. Basal leaves and buds accumulated the most raffinose. In evaluating responses to SD, experiments were conducted in greenhouse using ‘Cabernet Franc’, ‘Couderc 3309’ and ‘Concord’ cultivars. Shoot growth slowed under SD photoperiod in all cultivars. There was also increased periderm formation, endodormancy induction and increased FT. ‘Concord’ was the first cultivar to initiate these changes in response to SD followed by ‘Couderc-3309’ then ‘Cabernet franc’. Under LD, raffinose concentrations were low and similar among cultivars, whereas under SD, raffinose accumulation was higher and cold-tolerant cultivars ‘Couderc-3309’ and ‘Concord’ accumulated higher concentrations than the cold-sensitive cultivar ‘Cabernet franc’. In the field study we evaluated ‘Couderc 3309’, ‘Concord’ and ‘Cabernet Franc’ and characterized changes in FT and soluble sugar concentrations during the dormant season for two years. Generally basal buds were more FT than middle buds, which were more tolerant than apical buds. The cold-tolerant cultivars ‘Couderc 3309’ and ‘Concord’ had the lowest LT50 compared to the cold-sensitive ‘Cabernet Franc’. There was variation in leaf sugar concentration but leaf raffinose content showed cultivar dependent response associated with early acclimation. Cold-tolerant cultivars showed early responses to SD before low temperatures. In buds, among all sugars, fructose, glucose, sucrose, raffinose and stachyose concentrations had strong correlations with LT50. Basal buds accumulated the most raffinose. Raffinose accumulation was also two to three times iii higher in the cold-tolerant cultivars than in cold-sensitive. These results suggest that raffinose might be an early step in CA that coincides with early development of FT and can be used as a screening tool for breeding cold-tolerant genotypes and a target for improving FT via gene transfer. iv Dedication This dissertation is dedicated to my husband, Horatio Grant and my daughter, Breanne Grant v Acknowledgments First I must thank all my advisory committee members, Drs. Imed Dami, Rebecca Lamb, Jim Metzger, Ray Miller and Eric Stockinger, they have provided collaboration in designing experiments, interpreting data and also valued suggestions for this dissertation and publication. I would also like to thank my lab mates Miss Patricia Chalfant and Mr. Y. Zhang for assistance in collecting samples and data for my experiments. I am grateful to Dr. L. Phelan for his help with the GC machine; he has provided invaluable information that has helped me in completing this dissertation. I also thank Dr Taehyun Ji, Mr. David Scurlock, Mr. Bruce Williams and Miss Kesia Hartzler for helping with lab techniques, managing greenhouse plants and/or managing the vineyard in Wooster. I am thankful for the support from many undergraduate interns who helped with my experiments and I also appreciate all the help that I have received from the HCS faculty, staff and students as well as other facility staff at OARDC. This dissertation was also partially funded by SEEDS, a competitive grants program from The OARDC and the Lonz foundation. I am very grateful for the support my husband Horatio Grant has provided, his love has encouraged me to be the best I can be. I will forever be indebted to my daughter Breanne Grant for the joy and inspiration she gave me as I completed my PhD. vi Vita 1998 ...............................................................B.S. Biological Sciences, Northern Caribbean University 2007................................................................M.S. Biological Sciences Walla Walla College 2007 to present ..............................................Graduate Research Associate, Department Horticulture and Crop Science, The Ohio State University Publications Grant, T. N., Dami, I.E., Ji, T., Scurlock, D., and Streeter, J. 2009. Leaf raffinose is an early cold acclimation response in various Vitis genotypes. Can. J. Plant Sci. 89:961-968. Fields of Study Major Field: Horticulture and Crop Science vii Table of Contents Abstract ............................................................................................................................... ii Dedication ........................................................................................................................... v Acknowledgments.............................................................................................................. vi Vita .................................................................................................................................... vii Publications ....................................................................................................................... vii Fields of Study .................................................................................................................. vii Table of Contents ............................................................................................................. viii List of Tables ...................................................................................................................... x List of Figures .................................................................................................................. xiii Chapter 1: Introduction ...................................................................................................... 1 Chapter 2: Variation in leaf and bud soluble sugar concentration among Vitis genotypes grown under two temperature regimes ............................................................................. 23 Chapter3: Responses of greenhouse-grown Vitis genotypes to photoperiod regimes ...... 45 Chapter 4: Seasonal changes of freezing tolerance and soluble sugars among field-grown Vitis genotypes .................................................................................................................. 88 viii Chapter 5 Cold and short-day acclimation and how they affect grape genotypes of contrasting freezing tolerance ......................................................................................... 133 Bibliography ................................................................................................................... 140 Appendix A: GC Chromatograms .................................................................................. 148 Appendix B: LTE of grapevine Basal buds .................................................................... 151 ix List of Tables Table 2.1. Soluble sugar concentration (mg g-1 dry wt) in leaves of non-acclimated and acclimated Vitis vinifera, ‘Cabernet franc’, V. labruscana ‘Concord’, V. spp, ‘Frontenac’, ‘Traminette’ and ‘Seyval’ and V.riparia x V. rupestris ‘C-3309’ grapevines …………39 Table 2.2. Soluble sugar concentration (mg g-1 dry wt) in leaves and buds of non- acclimated and acclimated cold sensitive Vitis vinifera, ‘Cabernet Franc’ and cold hardy V. riparia x V. rupestris, ‘C-3309’grapevines…………………………………………...40 Table 2.3. Soluble sugar concentration (mg g-1 dry wt) in leaves of acclimated and non- acclimated Vitis vinifera, ‘Cabernet Franc’ at different stages of development z………..41 Table 3.1. Shoot length, number of nodes and periderm formation for Cabernet franc (CF), Couderc