engleski

Detection of factor V Leiden by PCR-SSCP using GMA (TM) precast elchrom scientific gels

Genetic abnormalities in hemostatic proteins associated with hypercoagulability are an important hereditary risk factor for venous thrombosis. Several genetic mutations that cause hereditary disorders predisposing to thrombosis have been described, point mutation in the coagulation Factor V gene (FV:R506Q), called Factor V Leiden, being the most common of them. Here we report on a new inexpensive and simple PCR-SSCP based method for detection of this genetic abnormality. Study population consisted of 150 subjects whose Factor V genotype was previously determined by PCR-RFLP method using the Mnl I restriction endonuclease. A 223bp fragment containing the G1692-A (Arg 506-Gln) polymorphic site in exon 10 of the Factor V gene was amplified, denatured and run overnight on the commercially available GMA™ gels for SSCP. PCR-SSCP analysis showed reproducible and uniform band patterns for FV mutant and wild type alleles. Furthermore, PCR-SSCP results were consistent with those obtained by PCR-RFLP analysis (100%). The described PCR-SSCP procedure is reliable, time saving and cost-effective. The method may be considered as a potentially powerful new tool in the routine detection of Factor V Leiden.

Factor V Leiden; PCR-SSCP; venous thrombosis

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