title: Polymerase chain reaction based genotyping for characterization of SLA-DQB and SLA-DRB alleles in domestic pigs.
abstract: Molecular genotyping of swine major histocompatibility complex SLA-DQB and SLA-DRB genes using polymerase chain reaction (PCR)-based amplification is described. Locus-specific oligonucleotide primers were designed for the analysis of expressed SLA genes by reverse transcription-polymerase chain reaction (RT-PCR). RT-PCR products were sequenced, and the information gained was used to design primers for PCR genotyping of the exon 2 (beta 1) region from genomic DNA templates. A single segregating amplification product was detected for both DQB and DRB in all animals. PCR products were digested with restriction enzymes. Seven SLA-DQB PCR-restriction fragment length polymorphism (RFLP) pattern types were observed for both HaeIII and RsaI that defined 14 SLA-DQB alleles. A total of seven SLA-DRB PCR-RFLP pattern types were defined using MspI (3 RFLP pattern types) and RsaI (6 RFLP pattern types). In order to demonstrate their universal utility, the primers were tested on genomic DNA samples from 10 different swine breeds. No breed-specific alleles were observed. These results show that locus-specific oligonucleotide primers and RFLP analysis provide a simple and rapid method for genotyping expressed SLA-DQB and SLA-DRB from genomic DNA.
authors.affiliation: Department of Animal Sciences, University of Illinois at Urbana-Champaign 61801, USA.
authors.forename: Y C
authors.initials: YC
authors.lastname: Shia
authors.forename: M
authors.initials: M
authors.lastname: Bradshaw
authors.forename: M S
authors.initials: MS
authors.lastname: Rutherford
authors.forename: H A
authors.initials: HA
authors.lastname: Lewin
authors.forename: L B
authors.initials: LB
authors.lastname: Schook
mesh_terms: D000483:Alleles; D000818:Animals; D001483:Base Sequence; D004247:DNA; D017931:DNA Primers; D005091:Exons; D005838:Genotype; D006649:Histocompatibility Antigens / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012150:Polymorphism, Restriction Fragment Length; D012194:RNA-Directed DNA Polymerase; D012689:Sequence Homology, Nucleic Acid; D013552:Swine / Q000235:genetics* / Q000276:immunology
chemical_list: D017931:DNA Primers; D006649:Histocompatibility Antigens; D004247:DNA; D012194:RNA-Directed DNA Polymerase