MAPPING OF MONOGENIC AND QUANTITATIVE TRAIT LOCI USING A WHOLE GENOME SCAN APPROACH AND SINGLE NUCLEOTIDE POLYMORPHISM PLATFORMS BY ALYSTA D. MARKEY DISSERTATION Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Animal Sciences in the Graduate College of the University of Illinois at Urbana-Champaign, 2013 Urbana, Illinois Doctoral Committee: Professor Jonathan Beever, Chair, Director of Research Professor Sandra Rodriguez-Zas Clinical Associate Professor Clifford Shipley Assistant Professor Anna Dilger Abstract The release of the bovine genome sequence in 2004 opened the door for the development of high density single nucleotide polymorphism (SNP) panels that can be used for linkage disequilibrium mapping of traits in cattle populations. The BovineSNP50 Beadchip, containing 54,001 SNP markers, was released in 2008 and the recently released BovineHD Beadchip contains 777,000 SNP markers. These SNP platforms were utilized to map both monogenic and quantitative traits to finite genomic regions in beef cattle populations. The recessive defect, hypotrichosis is an autosomal recessive form of hairlessness that affects Hereford cattle. A whole-genome association analysis was conducted using BovineSNP50 BeadChip to map the hypotrichosis locus to a chromosomal region. Significant association was detected between the hypotrichosis phenotype and a locus on bovine chromosome 5 (BTA5) and homozygosity analysis localized the associated region to between 29 and 32 Mb. Sequencing of six candidate genes in the region revealed an eight base pair deletion in exon 1 of the bovine KRT71 gene that was consistent with putative genotypic status within archived samples. The deletion mutation is predicted to result in a frameshift and early truncation of the K71 protein. A DNA based diagnostic was developed that will permit animals to be screened for the mutation and thus allow animal breeders to make informed mating decisions and decrease the incidence of hypotrichosis in the Hereford breed. Genotype imputation from the BovineSNP50 to the high density BovineHD platform was conducted to provide a denser marker panel to carry out whole genome association mapping and allow for more refinement of genomic regions associated with traits. The imputed genotypes were used in a whole genome scan in order to discover quantitative trait loci that may be influencing growth, carcass and meat quality traits in an US Simmental- Angus population. Association was detected with the traits birth weight, back fat, yield ii grade, ribeye area and marbling. There were 81 SNP associations with birth weight that were sequestered into 10 genomic regions. The carcass trait, back fat exhibited association with 209 SNPs in 46 genomic regions and yield grade displayed association with 172 SNPs in 69 genomic areas while there were four SNP associations with ribeye area, representing one genomic region. There were 127 SNP associations with the meat quality trait, marbling, that represented 32 genomic regions. These regions are ideal targets for development of markers for use in marker assisted selection and are also excellent regions to investigate with further fine mapping and discovery of causal variants underlying the quantitative trait loci. The utilization of genotype imputation from the BovineSNP50 to the high density 770K platform allowed localization of QTL to refined regions of the genome, often eliminating the need for further fine mapping efforts prior to targeted re-sequencing. The whole genome scan using imputed genotypes revealed a locus on BTA 6 from 37 to 42 Mb that was associated with back fat, marbling and birth weight in an US Simmental- Angus population. The region on BTA6 was evaluated in an attempt to further refine the candidate locus interval by implementing haplotype analysis with the imputed SNP markers that were used the whole genome association analysis. Two regions within the 5 Mb interval were associated with the phenotypes; the first was near 38.83 Mb that was associated with both back fat and birth weight. The second region near 39.27 Mb was associated with back fat, marbling and birth weight. The first region, near 38.83 Mb, corresponds to the LCORL-NCAPG locus that has been implicated in body size in many species as well as contributing to growth and carcass phenotypes in cattle. Re-sequencing of the LCORL-NCAPG locus in animals of know haplotype status revealed several polymorphisms that were consistent with haplotype status. This research contributes to the mounting evidence implicating the LCORL-NCAPG region as influencing growth, carcass and meat quality traits in cattle. iii Acknowledgements I have been blessed to have a great group of people behind me during my entire academic career and I am very thankful for each and every one of you. First of all I would like to thank my parents for their love and support not only in my academic pursuits but throughout my life. You have always been there for me and pushed me to excel while never expecting too much or being unrealistic. You raised me to appreciate the little things, to always respect others, to work hard and always do my best and for that I will be forever grateful. And to Jason – thank you for your love and support throughout this entire endeavor. I could not have done this without your understanding and desire for me to finish although I know it took me longer than either of us would have preferred. And yes, thank you for asking about my day and listening patiently even though I could see your eyes glaze over as I talked. I really appreciate that you care enough to ask! I would like to thank my lab-mates and good friends Erin Wagner, Brandy Marron, Kristen Walker and Stacey Meyers for everything they have done for me. I am truly blessed to have had such a great group of colleagues who all become more than just co-workers. I appreciate all the assistance with projects, moral support when things went awry, shared “wohoos” when things went right, and always having your support no matter what was happening. I am very happy that I have been able to work with group of people who had the ability to pull together and work as a cohesive team. I am so thankful to have had such a great work environment and without you all I am pretty certain I would have lost my sanity! We have shared many good times over the last few years and I will miss my daily interactions with each of you! I would like to thank “boss” for everything; both things that fell under official duties as well as those things in the unofficial category. I am thankful for all the guidance in research and teaching I truly appreciate your patience, constant availability and the fact that you really do care about each and every one of us. Although we all seem to be iv exasperated when you “show us how it’s done” it is nice to have a boss who can truly guide us, teach us, trouble shoot with us and who not only has the ability to work side by side with us but is always willing to do so. I am also grateful for all things you did that were not required of an advisor - lab lunches and birthday cakes, Christmas parties and cookouts, days on the Beever ranch working cows and the times that we simply sat around and had enjoyable conversations. All of these things are going to be hard to find again in future supervisors and I am truly thankful that I have had this good of an experience at least once! And lastly I would like to thank my committee members Sandra Rodriguez-Zas, Clifford Shipley and Anna Dilger. I appreciate the guidance from each of you and the contributions that you made to my thesis research and academic career at the University of Illinois. v Table of Contents List of Figures.........................................................................................................vii List of Tables..........................................................................................................viii Chapter 1. Literature Review ....................................................................................1 Introduction ....................................................................................................1 Current status of Bovine Genome Technologies .....................................................2 Trait Mapping Approaches ..................................................................................4 Selected Review of Trait Mapping ........................................................................9 Genotype Imputation.........................................................................................15 Chapter 2. A deletion mutation in KRT71 is associated with congenital hypotrichosis in.....17 Hereford cattle. Abstract ..........................................................................................................17 Background......................................................................................................18 Results and discussion .......................................................................................20 Conclusion .......................................................................................................29 Methods ..........................................................................................................30 Figures ............................................................................................................35 Chapter 3. A genome wide association analysis for growth, carcass and meat quality .......41 traits in cattle Abstract ..........................................................................................................41