P-59: Comparison of Two Methods for Screening of the MTHFR C677T Mutation in Patients with Hypercoagulable Defects


Poursadegh Zonouzi A *, Sakhinia E , karimi ansari N , Firoozi A ,

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Objective: Methylenetetrahydrofolate reductase (MTHFR) gene encodes a key enzyme in the homocysteine metabolism. The common mutation in MTHFR gene is C677T in the exon 4 which converts an alanine to a valine at codon 222 in the N-terminal catalytic domain of the protein related with a decreased activity of enzyme resulting in accumulation of homocysteine in the circulation. Previous studies have been suggested that a high concentration of blood homocysteine is a major risk factor for thrombophilic diseases. Materials and Methods: Most researchers generally use from PCR-RFLP method with the Hinf1 enzyme to identification MTHFR C677T mutation. We have focused to study the concordance between the PCR-RFLP using enzyme Hinf1 and ARMS-PCR methods to screening of C677T mutation among 225 people who were suffering from hypercoagulable defects. The genomic DNA extracted from peripheral blood by standard salting-out protocol was used to compare these techniques. To perform PCR-RFLP, the amplified DNA has been digested by Hinf1 enzyme and then, the fragments were analyzed by electrophoresis on polyacrylamide gel and stained with silver nitrate. On other hand, extracted DNA has been also amplified using other primers to perform ARMS-PCR and the PCR products were electrophoresed on 2% agarose gel and stained with ethidum bromaid. Results: Our results indicated that the PCR-RFLP is concordant with ARMS-PCR to screening of C677T mutation. Conclusion: Our results indicated that the PCR-RFLP is concordant with ARMS-PCR to screening of C677T mutation, suggesting that ARMS-PCR could be used as an alternative method for identification of this mutation because of ARMS-PCR is time and cost consuming method