title: A sensitive method for detecting variation in copy numbers of duplicated genes.
abstract: Gene duplications are common in the vertebrate genome, and duplicated loci often show a variation in copy number that may have important phenotypic effects. Here we describe a powerful method for quantification of duplicated copies based on pyrosequencing. A reliable quantification was obtained by amplification of the duplication break-point and a corresponding nonduplicated sequence in a competitive PCR assay. A comparison with an independent method for quantification based on the Invader technology revealed an excellent correlation between the two methods. The pyrosequencing-based method was evaluated by analyzing variation in copy number at the duplicated KIT/Dominant white locus in pigs. We were able to distinguish haplotypes at this locus by combining the duplication breakpoint test with a diagnostic test for a functionally important splice mutation in the duplicated gene. An extensive allelic variation, including the presence of a new allele carrying a single KIT copy expected to encode a truncated KIT receptor, was revealed when analyzing white pigs from commercial lines.
authors.affiliation: Department of Animal Breeding and Genetics, Swedish University of Agricultural Sciences, Uppsala, Sweden.
authors.forename: Gerli
authors.initials: G
authors.lastname: Pielberg
authors.forename: Andy E
authors.initials: AE
authors.lastname: Day
authors.forename: Graham S
authors.initials: GS
authors.lastname: Plastow
authors.forename: Leif
authors.initials: L
authors.lastname: Andersson
mesh_terms: D000465:Algorithms; D000483:Alleles; D000818:Animals; D005260:Female; D018628:Gene Dosage; D005799:Genes, Dominant / Q000235:genetics; D020131:Genes, Duplicate / Q000235:genetics*; D014644:Genetic Variation / Q000235:genetics*; D005828:Genetics, Population / Q000379:methods; D005838:Genotype; D006200:Hair Color / Q000235:genetics; D008297:Male; D010375:Pedigree; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods / Q000706:statistics & numerical data; D013552:Swine
chemical_list: D019009:Proto-Oncogene Proteins c-kit