A genotyping method for VKORC1 1173C > T by Pyrosequencing technology.

ABSTRACT

Vitamin K epoxide reductase complex subunit 1 (VKORC1) is the site of inhibition by warfarin and other antivitamin K drugs during oral anticoagulant therapy. The SNP rs9934438 in intron 1 of VKORC1 (c.173+1000C > T or 1173C > T) discriminating the VKORC1*2 haplotype is associated with low warfarin dose requirement and unstable prothrombin time - international normalized ratio. To genotype this SNP, we have developed a rapid method using Pyrosequencing technology. The proposed method takes a post-PCR sample preparation of less than 1 h and a DNA sequencing time of less than 15 min to genotype 96 samples. The current method was compared with a dHPLC method that we reported previously. Genotype frequencies at VKORC1 1173C>T for our Swedish population were 38% wild-type, 40% heterozygote and 22% homozygote. The frequency of the T-allele was 0.42, which exactly matches the frequency previously reported for Germans. The current method can be used to determine whether patients initiating warfarin therapy are carriers of SNP 1173 C>T that is strongly associated with low warfarin dose requirement.