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2012 | 68 | 09 |
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Phylogenetic analysis of steroid 21-hydroxylase (CYP21) of the wolf and selected breeds of dogs in connection with the role of this enzyme in the pathogenesis of many diseases in the Canidae family

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The aim of this study was to determine the nucleotide sequence of the CYP21 gene in the wolf and representatives of five breeds of dogs (selected according to a classification by Parker et al, 2004) in connection with the key role of the product of this gene in the genesis of many diseases in dogs. Nuclear DNA of dogs was obtained from peripheral blood, and the wolf’s DNA was isolated from muscle tissue. The amplification of the 21HS gene was carried out in 10 fragments under standardized PCR conditions. The reaction products were sequenced. The sequence of amino acids in the protein was determined on the basis of the nucleotide sequence. The sequences obtained in our study and those retrieved from the GeneBank database were compared with the Mafft program (15) and subjected to phylogenetic analysis with the MrBayes 3.2 program (35). We detected a total of nine SNP mutations in introns and exons. Furthermore, a deletion of two nucleotides, that differentiates the breeds, was detected in the promoter region. Only two differences between the dogs and the wolf were found in SNP: one in an exon and one in an intron. Genetic distance was determined between the selected breeds of dogs and between the wolf and the dogs of each breed. In addition, we estimated the evolutionary distances between amino acid sequences of the dog/wolf and homologous 21HS sequences of eight different vertebrate species obtained from GeneBank. It was shown that, among mammals, the amino acid sequence of the dog/wolf is the most similar to the sequence of the pig, and the least similar to that of the human. The sequences determined in this study may provide a reference point for the research on the CYP21 gene structure and expression in various tissues of dogs for therapeutic purposes.
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