Open AccessDeveloping and Evaluating the HRM Technique for Identifying Cytochrome P450 2D6 Polymorphisms
Author(s) -
Lu HsiuChin,
Chang YaSian,
Chang ChunChi,
Lin ChingHsiung,
Chang JanGowth
Publication year - 2015
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.21754
Subject(s) - genotype , cyp2d6 , high resolution melt , single nucleotide polymorphism , genetics , polymorphism (computer science) , biology , allele , microbiology and biotechnology , snp , genomic dna , gene
Background Cytochrome P450 2D6 is one of the important enzymes involved in the metabolism of many widely used drugs. Genetic polymorphisms of CYP2D6 can affect its activity. Therefore, an efficient method for identifying CYP2D6 polymorphisms is clinically important. Methods We developed a high‐resolution melting (HRM) analysis to investigate CYP2D6 polymorphisms. Genomic DNA was extracted from peripheral blood samples from 71 healthy individuals. All nine exons of the CYP2D6 gene were sequenced before screening by HRM analysis. This method can detect the most genotypes ( *1 , *2 , *4 , *10 , *14 , *21 *39 , and *41 ) of CYP2D6 in Chinese. Results All samples were successfully genotyped. The four most common mutant CYP2D6 alleles ( *1 , *2 , *10 , and *41 ) can be genotyped. The single nucleotides polymorphism (SNP) frequencies of 100C > T (rs1065852), 1039C > T (rs1081003), 1661G > C (rs1058164), 2663G > A (rs28371722), 2850C > T (rs16947), 2988G > A (rs28371725), 3181A > G, and 4180G > C (rs1135840) were 58%, 61%, 73%, 1%, 13%, 3%, 1%, 73%, respectively. We identified 100% of all heterozygotes without any errors. The two homozygous genotypes (1661G > C and 4180G > C) can be distinguished by mixing with a known genotype sample to generate an artificial heterozygote for HRM analysis. Therefore, all samples could be identified using our HRM method, and the results of HRM analysis are identical to those obtained by sequencing. Our method achieved 100% sensitivity, specificity, positive prediction value and negative prediction value. Conclusion HRM analysis is a nongel resolution method that is faster and less expensive than direct sequencing. Our study shows that it is an efficient tool for typing CYP2D6 polymorphisms.