Submitted to: Immunogenetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/12/2003
Publication Date: 6/20/2003
Citation: Martens, G.W., Lunney, J.K., Baker, J.E., Smith, D.M. 2003. Rapid assignment of swine leukocyte antigen (SLA) haplotypes n pedigreed herds using a polymerase chain reaction based assay. Immunogenetics. 55:395-401. Interpretive Summary: Many genes control how well a pig will grow and reproduce, and the quality of its carcass and meat. Independently, there are numerous genes which determine animal health. It has been known, and proven in many species, that there is one set of genes that are major determinants of disease resistance and some vaccine responses; these are the MHC [Major Histocompatibility Complex] genes, or for swine, the SLA [Swine Leukocyte Antigen] complex. Thus, as we pursue genes which regulate disease resistance and vaccination responses, it is essential that we can identify each of the alleles at the many SLA loci. Previously this involved a very labor intensive procedure of developing panels of serologic reagents and proving their specificity. Now that these genes have been sequenced more accurate molecular, PCR [Polymerase chain reaction] procedures using site specific primers [PCR-ssp] have become available. This paper represents the first development of these procedures for pigs. A combination of allele specific and positive control primers have resulted in a specific and robust PCR-ssp assays for assigning SLA haplotypes. With these assays we will be better prepared to assign the role of each SLA gene on immune and infectious disease responses.
Technical Abstract: We present a simple assay to determine the SLA haplotypes of animals within two experimental herds of MHC defined miniature pigs. The Yucatan Miniature Pigs have four founder haplotypes (w, x, y, z) and one recombinant haplotype (q). The NIH Miniature Pigs have three founder haplotypes (a, c, d) and two recombinant haplotypes (f, g). Since most crossovers occur between the class I and class II regions, for practical purposes, haplotypes can be assigned by typing one class I locus and one class II locus. We have previously characterized these seven founder haplotypes by sequencing the cDNA of three SLA class I loci, designated as SLA1, SLA3 and SLA-B, and four SLA class II loci, SLA-DQA1, DQB1, DRA1 and DRB1. Based on these sequences, allele specific primers were designed to amplify one MHC class I and one MHC class II gene for each haplotype and tested for specificity in homozygous and heterozygous animals. Positive control primers were also designed to amplify a portion of the E-selectin or alpha-actin gene and multiplexed with the allele specific primers to check for false negatives. This combination of allele specific and positive control primers produced specific and robust PCR-site specific primer (PCR-ssp) assays for assigning SLA haplotypes in the two herds.