SIMULTANEOUS POLYMERASE-CHAIN-REACTION RESTRICTION-FRAGMENT-LENGTH-POLYMORPHISM IDENTIFICATION OF THE FACTOR-V-LEIDEN ALLELE AND THE PROTHROMBIN 20210A MUTATION

A novel mutation in the 3' untranslated region of the prothrombin gene , prothrombin 20210A, recently has been identified. This mutation is a ssociated with increased serum prothrombin levels and an increased ris k for venous thromboembolism. Patients who carry a mutation in the fac tor V gene (factor V Leiden) have also been demonstrated to be at incr eased risk for venous thromboembolism, and previous studies have ident ified a population prevalence of approximately 5% to 10% for the facto r V Leiden allele. To simply and reliably identify patients who carry both genetic defects, a novel assay was developed that simultaneously determines the genotype of patients for the factor V Leiden allele and the prothrombin 20210A mutation. Representative samples (samples posi tive and negative for each mutation and a ''double mutant'') were then subjected to this single-tube genotyping assay. The results indicate that the simultaneous genotyping of these mutations will readily chara cterize the allelic status of patients for the two most frequent genet ic mutations in the coagulation cascade.