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Genomic Evolution of Vitis Vinifera – a Palaeogenomic Approach

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Genomic Evolution of Vitis Vinifera – a Palaeogenomic Approach FACULTY OF SCIENCE UNIVERSITY OF COPENHAGEN Genomic Evolution of Vitis vinifera Master’s thesis Insights into the history of viniculture from ancient DNA Eva Egelyng Sigsgaard Master thesis by Anne Kathrine Wiborg Runge Monitoring of marine fishes using environmental DNA from seawater samples Academic advisors: Post Doc. Nathan Wales. Professor Tom Gilbert Submitted 31.08.2015 Academic advisors: Postdoc Philip F. Thomsen and Ass. Prof. Peter R. Møller Submitted:! 01/08/14 ! 1! Institute name: Natural History Museum of Denmark Name of department: Centre for Geogenetics Author: Anne Kathrine Wiborg Runge Title: Genomic Evolution of Vitis vinifera – a palaeogenomic approach Subject description: This MSc thesis presents the results of the first in-solution target capture on ancient and historic Vitis vinifera subsp vinifera seeds and explores their potential in ancient DNA studies. An enrichment of up to 3.4 fold was observed in bioinformatically useful reads in post-capture libraries. Berry colour was inferred for 18 samples from the Roman and Medieval period. Furthermore a tentative assignment of sex genotype was made for seven berries. Evidence of a genetic displacement in cultivars through time was found at one site on the coast of the Mediterranean Basin, matching the changes in human habitation in the area. Finally, a provisional identification of two Pinot noir and two Chradonnay cultivars was made. In all, this study proves the effectiveness of target enrichment of ancient plants DNA, and demonstrates the advantages of using ancient and historic grape seeds to study domestication in plants. Academic advisors: Post Doc. Nathan Wales. Centre for GeoGenetics Natural History Museum of Denmark University of Copenhagen Professor Tom Gilbert Centre for GeoGenetics Natural History Museum of Denmark University of Copenhagen Submitted: 31st August 2015 Cover illustration: photo by Anne Kathrine Wiborg Runge, Montpellier, France (2015) ,,Grapes.“ ! ! 2! Table of contents PREFACE AND ACKNOWLEDGEMENTS!......................................................................................................!4! SUMMARY!..................................................................................................................................................................!5! INTRODUCTION!......................................................................................................................................................!6! A REVIEW OF THE HISTORY OF ANCIENT DNA RESEARCH!..............................................................................................!6! Introducing aDNA!.......................................................................................................................................................................!6! The methodological progress in aDNA research!...........................................................................................................!7! Ancient plants!.............................................................................................................................................................................!12! GRAPES!..........................................................................................................................................................................................!13! Domestication history!..............................................................................................................................................................!14! Domestication traits!..................................................................................................................................................................!15! METHODS!.................................................................................................................................................................!20! SAMPLES!.......................................................................................................................................................................................!20! EXTRACTION AND LIBRARY BUILD!......................................................................................................................................!20! CAPTURE!.......................................................................................................................................................................................!24! DATA ANALYSIS!.........................................................................................................................................................................!28! RESULTS!...................................................................................................................................................................!31! TARGET ENRICHMENT!..............................................................................................................................................................!31! SEX DETERMINATION!................................................................................................................................................................!33! BERRY COLOUR!..........................................................................................................................................................................!34! POPULATION STRUCTURE!.......................................................................................................................................................!37! DISCUSSION!.............................................................................................................................................................!39! TARGET ENRICHMENT!..............................................................................................................................................................!39! GRAPE SEX!....................................................................................................................................................................................!40! BERRY COLOUR!..........................................................................................................................................................................!42! POPULATION STRUCTURE!........................................................................................................................................................!43! CONCLUDING REMARKS!...........................................................................................................................................................!44! REFERENCES!..........................................................................................................................................................!45! SUPPLEMENTARY MATERIAL!.......................................................................................................................!51! SUPPLEMENTARY FIGURES!......................................................................................................................................................!51! ! ! 3! Preface and acknowledgements This M.Sc. thesis was conducted at the Centre for GeoGenetics, Natural History Museum of Copenhagen, University of Copenhagen, and was supervised by Nathan Wales and Tom Gilbert. I would like to thank Laurent Bouby for providing the grape seeds used in the experiments described below, and Roberto Bacilieri and the GrapeReSeq project for providing the SNPs used in the capture experiment, access to the their database on modern cultivars, and valuable discussion on every aspect of grapes. In addition, their hospitality during my visit in Montpellier was greatly appreciated. I would like to thank my supervisors for being dedicated and inspiring, for their help and guidance, and for constructive discussions during the process. I would like to express my gratitude to Jazmín Ramos Madrigal for the help she gave me with the bioinformatics, her patience in teaching me how to work in the terminal, and the tremendous amount of work she put into the analytical phase. I would also like to thank Jose Alfredo Samaniego for designing the RNA baits used in the capture experiment, and everybody at the Centre for GeoGenetics and the Danish National High-throughput DNA Sequencing Centre for their help and guidance. ! ! 4! Summary This MSc thesis presents the results of the first in-solution target capture on ancient and historic Vitis vinifera subsp vinifera seeds, and explores their potential in ancient DNA studies. An enrichment of up to 3.4 fold was observed in bioinformatically useful reads in post-capture libraries. Berry colour was inferred for 18 samples from the Roman and Medieval period. Furthermore a tentative assignment of sex genotype was made for seven berries. Evidence of a genetic displacement in cultivars over a 1000 year period was found, which matches the changes in human habitation in the area. Finally, a provisional identification of two Pinot blanc and two Chardonnay cultivars was made. In all, this study proves the effectiveness of target capture in enriching for ancient plant DNA, and demonstrates the advantages of using ancient and historic grape seeds to study domestication in plants. ! ! 5! Introduction This study has four main goals, 1) to determine the benefits of using an array-based in-solution capture protocol on waterlogged ancient and historic Vitis seeds, 2) to investigate the possibility of determining sex genotypes in ancient and historic grape pips, and thereby
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