Factor V Leiden Detection by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism with Mutagenic Primers in a Multiplex Reaction with Pro G20210A--A Novel Technique

Abstract

Factor V Leiden (FVL) R506Q and Prothrombin G20210A are clinically important genetic mutations associated with increased susceptibility to venous thrombosis. The objective of our study was to design a polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) reaction that allows simultaneous detection of these two mutations. The reaction can be used in routine diagnostic settings. We have analysed 4504 alleles for each mutation with a mutagenic primer-based PCR system with a low failure rate. The system eliminates the false positive FVL G1691A results associated with other PCR/RFLP caused by rare confounding mutations adjacent to restriction endonuclease recognition sites. This multiplex PCR/RFLP reaction is rapid, robust and dependable.

The Predictability of Factor V Leiden (FV:Q506) Gene Mutation via Clotting-Based Diagnosis of Activated Protein C Resistance

“…Generally, two major approaches have been applied for the detection of APCR and factor V Leiden mutation at the hemostasis laboratories. A wide variety of functional clotting-based methods were developed for the investigation of APCR (10-26) and specific DNA-based structural gene analysis has been equipped to search FV:Q 506 Leiden mutation (3,4,9,(27)(28)(29)(30)(31).Effective optimal screening strategy for the laboratory diagnosis of thrombophilia is a current questionable puzzle in clinical medicine (32)(33)(34)(35)(36). That obstacle includes the recognition of APCR due to FV:Q 506 , because this kind of 265…”

“…Generally, two major approaches have been applied for the detection of APCR and factor V Leiden mutation at the hemostasis laboratories. A wide variety of functional clotting-based methods were developed for the investigation of APCR (10-26) and specific DNA-based structural gene analysis has been equipped to search FV:Q 506 Leiden mutation (3,4,9,(27)(28)(29)(30)(31).…”