Single-strand conformation polymorphism (SSCP) assays for major histocompatibility complex B genotyping in chickens.

نویسندگان

  • R M Goto
  • M Afanassieff
  • J Ha
  • G M Iglesias
  • S J Ewald
  • W E Briles
  • M M Miller
چکیده

We have developed a DNA-based method for defining MHC B system genotypes in chickens. Genotyping by this method requires neither prior determination of allele-specific differences in nucleotide sequence nor the preparation of haplotype-specific alloantisera. Allelic differences at chicken B-F (class I) and B-L (class II) loci are detected in PCR single-strand conformation polymorphism (SSCP) assays. PCR primer pairs were designed to hybridize specifically with conserved sequences surrounding hypervariable regions within the two class I and two class I loci of the B-complex and used to generate DNA fragments that are heat- and formamide-denatured and then analyzed on nondenaturing polyacrylamide gels. PCR primer pairs were tested for the capacity to produce SSCP patterns allowing the seven B haplotypes in the MHC B congenic lines, and seven B haplotypes known to be segregating in two commercial broiler breeder lines to be distinguished. Primer pairs were further evaluated for their capacity to reveal the segregation of B haplotypes in a fully pedigreed family and in a closed population. Concordance was found between SSCP patterns and previously assigned MHC types. B-F and B-L SSCP patterns segregated in linkage as expected for these closely linked loci. We conclude that this method is valuable for defining MHC B haplotypes and for detecting potential recombinant haplotypes especially when used in combination with B-G (class IV) typing by restriction fragment pattern.

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عنوان ژورنال:
  • Poultry science

دوره 81 12  شماره 

صفحات  -

تاریخ انتشار 2002