Research Article RFLP-PCR is more efficient than ARMS-PCR for identifying CYP2C19*2 polymorphism in atherosclerotic patients

Abstract

The CYP2C19*2 polymorphism is the result of a point mutation leading to an alternative splicing defect, generating an enzyme with a reduced function. The enzyme CYP2C19 is responsible for the activation of the prodrug Clopidogrel and the loss of its function leads to an increase in platelet aggregation. We compared the ARMS-PCR (Amplification Refraction Mutation System) and RFLP-PCR (Restriction Fragment Length Polymorphism) to determine which of the techniques was more efficient for identifying the CYP2C19*2 polymorphism. We analyzed 200 samples from patients with atherosclerosis and 100 samples from a control group, free from the disease. The polymorphic allele was not observed in any of the samples analyzed by the ARMS-PCR technique, but was present in more than 70% of the samples when analyzed by RFLP-PCR. Thus, RFLP-PCR generated more satisfactory results for the analysis of CYP2C19*2 polymorphism, and it should be used, despite the lower cost of ARMS-PCR, as the latter did not detect this polymorphism.

Exploring the Potential and Limitations of PCR-RFLP and PCR-SSCP for SNP Detection: A Review

“…The apparent superiority of PCR-RFLP was observed regarding its broad utilization in several aspects of medical human genetics, such as the diagnosis of carcinogenesis, parasitic infection, gastritis, urinary tract infection, arthrosclerosis, infertility, and blood grouping. [40][41][42][43][44][45][46][47][48] This higher reliability on PCR-RFLP may be attributed to its simplicity compared with PCR-SSCP, because of which it has been used in previous SNP-specified applications. In contrast to medical applications, PCR-RFLP has not exhibited superiority compared with PCR-SSCP.…”

Comment: “Evaluation of the Association of Omentin 1 rs2274907 A>T and rs2274908 G>A Gene Polymorphisms with Coronary Artery Disease in Indian Population: A Case Control Study”

“…As in the ARMS system, the design of compatible primers for simultaneous amplification is often challenging, and it has been shown to increase the probability of errors. A recent study [ 5 ] highlighted the inefficacy of the ARMS-PCR, where this technique failed to identify the amplification of the polymorphic allele. Some of the most common causes for the inefficacy of the technique include hairpin loop formation, annealing with the other primer (primer dimer) and dramatic difference in annealing temperature of each primer.…”