Real-Time PCR Assay for Rapid Detection of GSTM1 Polymorphism in Nasopharyngeal Carcinoma Patients

Nasopharyngeal carcinoma (NPC) is a common public health problem in Thailand. Glutathione S-transferaseM1 gene deletion (GSTM1 null genotype) carriers have been reported to be at increased risk and therefore thisparameter is a potential marker for screening of NPC high-risk individuals. However, the conventional polymerasechain reaction (C-PCR) assay commonly used for GSTM1 null genotype detection is not suitable for mass screeningsince it is inconvenient, time consuming and unsafe due to the use of a toxic chemical. Currently, real-time PCR(R-PCR) assay is recommended for quicker and safer detection of various genetic polymorphisms. The aim ofthis study was to develop a SYBR green I R-PCR assay combined with melting curve analysis for GSTM1polymorphism detection in Thai NPC patients. The results were compared to those from the C-PCR assay usingDNA samples from peripheral blood leukocytes of 120 Thai NPC patients. The frequencies of GSTM1polymorphism detected by the R-PCR and the C-PCR were the same. Forty-eight individuals that were GSTM1+in the R-PCR assay showed 2 peaks with melting points of 82.5˚C and 87.5˚C that correlated with the appearanceof 2 DNA bands in the C-PCR assay (i.e., one for GSTM1 at 215 base pairs (bp) and one for β-globin at 268 bp).By contrast, 72 individuals that were GSTM1- in the R-PCR assay showed 1 peak with a melting point of 87.5˚Cthat correlated with the appearance of 1 DNA band for β-globin at 268 bp in the C-PCR assay. The R-PCR assayusing SYBR Green I and melting curve analysis for GSTM1 polymorphism detection was as reliable as the CPCRassay but was quicker and safer and more amenable to large scale screening in Thai NPC cases.