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-transferase M1 gene deletion (GSTM1 null genotype) carriers have been reported to be at increased risk and therefore this parameter is a potential marker for screening of NPC high-risk individuals. However, the conventional polymerase chain reaction (C-PCR) assay commonly used for GSTM1 null genotype detection is not suitable for mass screening since 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 of this study was to develop a SYBR green I R-PCR assay combined with melting curve analysis for GSTM1 polymorphism detection in Thai NPC patients. The results were compared to those from the C-PCR assay using DNA samples from peripheral blood leukocytes of 120 Thai NPC patients. The frequencies of GSTM1 polymorphism 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 and 87.5 that correlated with the appearance of 2 DNA bands in the C-PCR assay (i.e., one for GSTM1 at 215 base pairs (bp) and one for ?-globin at 268 bp). By contrast, 72 individuals that were GSTM1?- in the R-PCR assay showed 1 peak with a melting point of 87.5C that correlated with the appearance of 1 DNA band for -globin at 268 bp in the C-PCR assay. The R-PCR assay using SYBR Green I and melting curve analysis for GSTM1 polymorphism detection was as reliable as C-PCR assay but was quicker and safer and more amenable to large scale screening in Thai NPC cases.
Tiwawech
Danai
D
Research Division, National Cancer Institute, Bangkok 10400, Thailand. tdanai@hotmail.com
Chindavijak
Somjin
S
Karalak
Anant
A
Ishida
Takafumi
T
D020022:Genetic Predisposition to Disease; D005982:Glutathione Transferase / Q000235:genetics*; D006801:Humans; D009303:Nasopharyngeal Neoplasms / Q000235:genetics* / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic / Q000235:genetics*; D013785:Thailand / Q000453:epidemiology
D005982:Glutathione Transferase