IDENTIFYING THE C677T POLYMORPHISM OF MTHFR GENE BY PCR-RFLP TECHNIQUE

Abstract

Background: The C677T polymorphism of MTHFR gene is a risk factor of many diseases. This study is aimed at: (1) Improving a PCR-RFLP process with the own designed primers to identify the C677T polymorphism of MTHFR gene. (2) Evaluating the prevalence of the C677T polymorphism of MTHFR gene in volunteer group. Materials and method: DNA samples was extracted from peripheral blood of 60 volunteers. Designing primers by using FastPCR software, then improving PCR technique. Standardizing the optimal conditions of restriction digest by HinfI. Confirming the results of polymorphism by DNA sequencing technique. Results: We designed successfully primers to amplify fragment of MTHFR gene including C677T polymorphism and an obligatory restriction site of HinfI (as internal control). 0.5 µl of HinfI enzyme (10 U/µl) is enough for restriction digest. The MTHFR genotype frequencies were: 71.67 % (677CC); 25% (677CT); and 3.33 % (677TT). Conclusion: We standardized successfully PCR-RFLP technique to identifying C677T polymorphism of MTHFR gene. Keywords: C677T polymorphism, MTHFR gene, PCR-RFLP