University of Milan Department of Veterinary Medicine Ph.D. Course in Veterinary and Animal Sciences Class 29 CNV detection and association studies in the Brown Swiss cattle breed Raphaëlle T.M.M. Prinsen R10464 – R35 Tutor: Prof. Alessandro Bagnato Ph.D. Coordinator: Prof. Fulvio Gandolfi Academic Year: 2016 Academic year: 2016 CNV detection and association studies in the Brown Swiss cattle breed Raphaëlle T.M.M. Prinsen 2 To Andrea, my beloved husband, who gave me endless love, support, encouragement and endurance to complete this hard work. … and to our little baby: you are the most beautiful gift I could ever imagine. 3 Contents Sintesi – Italiano ............................................................................... 7 Capitolo 1 ................................................................................................................................ 7 Capitolo 2 ................................................................................................................................ 8 Capitolo 3 ................................................................................................................................ 9 Abstract – English ........................................................................... 10 Chapter 1 .............................................................................................................................. 10 Chapter 2 .............................................................................................................................. 11 Chapter 3 .............................................................................................................................. 12 Abbreviations ................................................................................. 13 General Introduction ...................................................................... 14 Copy Number Variants ................................................................................................... 14 CNVs identification and CNV studies........................................................................ 15 CNVs and Phenotypes ..................................................................................................... 17 CNVs and Genome-Wide Association Studies ....................................................... 18 Aims ....................................................................................................................................... 19 References ........................................................................................................................... 20 1. Identification and validation of copy number variants in Italian Brown Swiss dairy cattle using the Illumina Bovine SNP50 Beadchip 28 Abstract ................................................................................................................................ 28 Introduction ........................................................................................................................ 29 Material and Methods ..................................................................................................... 31 Sampling and genotyping ............................................................................................... 31 Editing data ........................................................................................................................... 32 Copy number variations detection .............................................................................. 32 PennCNV detection ............................................................................................................. 33 SVS7 detection ...................................................................................................................... 33 Copy number variation regions validation by quantitative polymerase chain reaction ....................................................................................................................... 34 Copy number variation regions annotation ........................................................... 34 Results and discussion ................................................................................................... 35 Copy number variations and copy number variation regions detection .. 35 Copy number variation regions validation by quantitative polymerase chain reaction ....................................................................................................................... 38 Copy number variation regions annotation ........................................................... 38 Conclusions ......................................................................................................................... 41 References ........................................................................................................................... 42 Supporting information ................................................................................................. 55 2. A genome-wide scan of copy number variants using high-density SNPs in Brown Swiss dairy cattle ................................................... 103 Abstract .............................................................................................................................. 103 Introduction ...................................................................................................................... 105 Material and methods ................................................................................................... 107 Sampling and genotyping ............................................................................................ 107 Quality assurance of CNV raw data ......................................................................... 107 Copy Number Variation detection ........................................................................... 108 CNVRs and consensus CNVRs definition ................................................................ 108 Consensus CNVR annotation ....................................................................................... 109 CNVR validation by qPCR ............................................................................................. 109 Results and discussion ................................................................................................. 110 Quality assurance of CNV Raw Data ....................................................................... 110 CNVs detection .................................................................................................................. 110 CNVRs and consensus CNVRs definition ................................................................ 112 Consensus CNVR annotation ....................................................................................... 115 CNVR validation by qPCR ............................................................................................. 117 Conclusion ......................................................................................................................... 118 References ......................................................................................................................... 118 Supporting information ............................................................................................... 128 3. A genome wide association study between CNVs and quantitative traits in Brown Swiss cattle ........................................................... 163 Abstract .............................................................................................................................. 164 Introduction ...................................................................................................................... 164 Material and methods ................................................................................................... 166 Sampling and genotyping ............................................................................................ 166 Phenotypes .......................................................................................................................... 166 CNV-association with productive, udder and health traits .......................... 167 CNVs annotation............................................................................................................... 167 Results and discussion ................................................................................................. 168 CNV-association and annotation results ............................................................... 168 Conclusion ......................................................................................................................... 180 Conflict of interest statement .................................................................................... 181 Acknowledgements ....................................................................................................... 182 References ......................................................................................................................... 182 5 General Discussion ....................................................................... 188 Extra – curricular activities ............................................................ 191 Conferences, congresses and workshops ............................................................. 191 Publications .....................................................................................................................