RADIATION HYBRID MAPPING OF AND IDENTIFICATION OF SINGLE NUCLEOTIDE POLYMORPHISMS IN PIG SEQUENCE TAGGED-SITES

 

 

Valencia D. Rilington1, Patrick J. Venta2, Donna Housley2, Nancy E. Raney1, Stephanie R. Wesolowski1, Christopher P. Wilkinson1 and Catherine W. Ernst1

1Department of Animal Science and 2Departments of Microbiology and Molecular Genetics, and Small Animal Clinical Sciences, Michigan State University, East Lansing MI 48824

 

Comparative mapping allows transfer of information from marker-rich genomes such as the human to species with limited mapping resources. Current pig genome maps contain relatively few Type I (gene) markers and mapping of such markers will clarify gene order and improve the utility of comparative maps. The objective of an ongoing project in our laboratory is to increase the density of the pig comparative map through the development and mapping of pig sequence tagged-sites (STS). The need for identifying additional polymorphism in Type I markers for genetic linkage mapping can be fulfilled by detection of single-nucleotide polymorphisms (SNPs). For SNP identification, we use a pool-and-sequence strategy in which a pool of DNA samples from different breeds is PCR-amplified and sequenced. SNPs are identified as ambiguous bands occurring at identical positions in a sequencing gel. Here we report the development of 133 new pig STS, the radiation hybrid (RH) mapping of 52 STS using the INRA-University of Minnesota porcine RH panel and the identification of 31 SNPs in 22 STS (1-5 SNPs per STS). PCR primers were designed using heterologous sequences and the identity of each STS was confirmed by DNA sequencing. Two-point analysis of RH data showed significant linkage of STS with markers on 12 different pig chromosomes. Genetic linkage analysis has been performed using SNP genotypes for 7 STS in the PiGMaP reference families. These STS include calcium channel voltage dependent L type alpha 1D subunit (CACNA1D, SSC13, nearest marker Sw864, LOD 7.41), corticotropin releasing hormone receptor 2 (CRHR2, SSC10, nearest marker S0120, LOD 5.48), dopamine beta hydroxylase (DBH, SSC1, nearest marker S0303, LOD 5.72), insulin-like growth factor binding protein-2 (IGFBP2, SSC15, nearest marker Sw936, LOD 4.8), 26S proteasome non-ATPase regulatory subunit 1 (PSMD1, SSC15, nearest marker Sw1119, LOD 5.12), solute carrier family 6 member 4 (SLC6A4, SSC12, nearest marker Sj018, LOD 3.91), and talin 1 (TLN1, SSC1, nearest marker S0113, LOD 6.73). Rapid genotyping assays for linkage mapping of additional SNPs are being developed. Results from this study will help to further improve the pig and human comparative map.

 

Keywords: pig, gene mapping, single nucleotide polymorphism