MHC class I typing in a songbird with numerous loci and high polymorphism using motif-specific PCR and DGGE.
The major histocompatibility complex (MHC) has a central role in the specific immune defence of vertebrates. Exon 3 of MHC class I genes encodes the domain that binds and presents peptides from pathogens that trigger immune reactions. Here we develop a fast population screening method for detecting genetic variation in the MHC class I genes of birds. We found evidence of at least 15 exon 3 sequences in the investigated great reed warbler individual. The organisation of the great reed warbler MHC class I genes suggested that a locus-specific screening protocol is impractical due to the high similarity between alleles across loci, including the introns flanking exon 3. Therefore, we used motif-specific PCR to amplify two subsets of alleles (exon 3 sequences) that were separated with by DGGE. The motif-specific primers amplify a substantial proportion of the transcribed class I alleles (2-12 alleles per individual) from as many as six class I loci. Although not exhaustive, this gives a reliable estimate of the class I variation. The method is highly repeatable and more sensitive in detecting genetic variation than the RFLP method. The motif-specific primers also allow us to avoid screening pseudogenes. In our study population of great reed warblers, we found a high level of genetic variation in MHC class I, and no less than 234 DGGE genotypes were detected among 248 screened individuals.
Westerdahl
H
H
Department of Animal Ecology, Ecology Building, Lund University, S-223 62 Lund, Sweden. Helena.Westerdahl@zooekol.lu.se
Wittzell
H
H
von Schantz
T
T
Bensch
S
S
D000483:Alleles; D000818:Animals; D017931:DNA Primers; D005091:Exons / Q000235:genetics*; D005260:Female; D005805:Genes, MHC Class I / Q000235:genetics*; D014644:Genetic Variation / Q000235:genetics*; D005828:Genetics, Population; D005838:Genotype; D008297:Male; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D020308:Songbirds / Q000235:genetics*
D017931:DNA Primers