Polymerase chain reaction-based genotyping of swine MHC DQB and DRB alleles
Shia, Yun-Chao
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Permalink
https://hdl.handle.net/2142/19201
Description
Title
Polymerase chain reaction-based genotyping of swine MHC DQB and DRB alleles
Author(s)
Shia, Yun-Chao
Issue Date
1993
Doctoral Committee Chair(s)
Schook, Lawrence B.
Department of Study
Animal Sciences
Discipline
Animal Sciences
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Genetics
Agriculture, Animal Pathology
Language
eng
Abstract
"Molecular genotyping the swine major histocompatibility complex DQB and DRB loci using the PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) analysis technique is described. SLA DQB and DRB locus-specific oligonucleotide primers were designed to analyze expressed genes using the reverse transcription-polymerase chain reaction (RT-PCR) method. Five SLA DQB alleles and three SLA DRB alleles were sequenced after oligonucleotide-directed amplification of mRNA. Sequence alignment studies of SLA DQB and DRB $\beta\sb1$ domain sequences provided a similarity index among seven SLA DQB alleles of 87%, and the similarity index among five SLA DRB alleles was 85%; the similarity index among the SLA DQB and DRB alleles was 61%. Using this sequence information, SLA DQB and DRB oligonucleotide primers were designed for PCR genotyping the SLA DQB and DRB exon 2 ($\beta\sb1$) region from genomic DNA. These oligonucleotide primers were tested on genomic DNA from ten different swine breeds (Meishan, Yorkshire, Hampshire, Duroc, Landrace, Berkshire, Chester White, Spotted Poland China, Poland China and Swedish Landrace) to confirm their ""universal"" utility. DNA products after genomic PCR amplifications were subjected to genotyping by restriction enzyme digestion and subsequently electrophoresed on polyacrylamide gels. Several SLA DQB PCR-RFLP pattern types were observed using HaeIII (7 RFLPs) and RsaI (7 RFLPs): SLA DRB PCR-RFLP pattern types using MspI (3 RFLPs) and RsaI (5 RFLPs) were also demonstrated. Sixteen SLA DQB alleles and seven SLA DRB alleles were defined. Thus, locus-specific oligonucleotide primers provide a simple and rapid method for analyzing SLA DQB and DRB from genomic DNA isolated from small blood samples."
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