Combination of multiplex PCR and DHPLC-based strategy for CYP2D6 genotyping scheme in Thais

Abstract

ObjectiveTo develop CYP2D6 genotyping scheme for accurate allele calling and reliable estimation of functional allele dosage in Thais. Design and methodsWe analyzed CYP2D6 copy numbers by pentaplex PCR coupled with semi-quantitative denaturing high performance liquid chromatography (DHPLC)-based technique. Ten common SNPs were genotyped from CYP2D6 gene product using single base extension (SBE) followed by DHPLC analysis. This detection scheme was compared with real-time PCR and conventional PCR-RFLP for cost-effectiveness. ResultsThe distribution of CYP2D6 gene copy numbers in our population ranged from zero (0.69%), one (7.99%), two (60.07%), three (28.13%) and four (3.13%). The most commonly detected SNPs were related to CYP2D6 * 10 haplotype. CYP2D6 * 36 in tandem with CYP2D6 * 10B is the major rearrangement type in Thais (18.75%). ConclusionsMultiplex PCR coupled with DHPLC-based strategy is convenient and reliable method for CYP2D6 genotyping offering sufficient allele coverage for Asians. Both cost and analytical time saving were shown and the method could potentially be modified to accommodate CYP2D6 genotyping in other ethnics. © 2011 The Canadian Society of Clinical Chemists.