Simultaneous genotyping of 11 non-synonymous SNPs in the 4 glutathione peroxidase genes using the multiplex single base extension method

Abstract

Background Glutathione peroxidase (Gpx) is one of the major antioxidant enzymes involved in scavenging hydrogen peroxide and lipid hydroperoxides produced during normal metabolism or after oxidative insult. In the Gpx genes, a number of single nucleotide polymorphisms (SNPs) have been identified and their associations with various diseases have been reported. In the present study, we used a multiplex single base extension (MSBE) technique to genotype multiple non-synonymous SNPs in the Gpx1, Gpx2 , Gpx3 and Gpx4 genes simultaneously. Methods Seven templates for the MSBE reaction, involving 11 SNPs corresponding to non-synonymous mutations were amplified by multiplex PCR. Availability of the MSBE method was validated by genotyping DNA from 91 Japanese, 91 German and 93 Xhosa healthy subjects. Results A simple and reproducible method for simultaneous genotyping of multiple SNPs in the Gpx genes was established. Of the 11 SNPs, only Gpx1 P200L was polymorphic in all three ethnic groups and the genotype distributions differed significantly among the three populations. On the other hand, little heterogeneity was observed for Gpx2 R146C, Gpx2 P126L, Gpx1 A194T and Gpx4 S2N, and no heterogeneity was observed for Gpx1 L6P, Gpx1 R5P, Gpx3 F128L, Gpx3 K144X, Gpx4 A88V and Gpx4 S227L. Conclusion The MSBE procedure described here was proven to be applicable for population studies. Application of this method will provide comprehensive information for studying the relationship between SNPs in the Gpx genes and risks for various diseases.