Detection of Arg72Pro polymorphism of the tumor suppressor gene (TP53) by a rapid one-step tetra-primer amplification refractory mutation system-PCR

Abstract

The Tumor Protein p53 gene (TP53), known as the guardian of the genome, encodes the p53 protein that plays an important role in cell cycle and in maintenance of the genome stability by preventing mutations. The transition from G to C at codon 72 of the TP53 gene represents a non-synonymous single nucleotide polymorphism (Arg72Pro) that has been extensively genotyped for association with a wide variety of cancers. A new method for Arg72Pro detection is described based on the tetra-primer amphfication refractory mutation system-polymerase chain reaction (ARMS-PCR), with a single PCR to discriminate both alleles. Two primers that amplify a common amplicon independently of the allele considered are combined in a single PCR reaction with two specific primers, differing in the 3' base. In the presence of Pro/Pro or Arg/Arg, amplification occurs both in the general amplicon and in the specific allele; in the presence of Arg/Pro three different amplicons are produced. Validation by PCR-RFLP revealed 100% accordance for genotype assignment. The method was successfully applied for genotype determination of the Arg72Pro in a novel population from Aleppo, Syria, with 51% homozygous for the Arg allele and 12% homozygous for the Pro Allele. © 2011 Academic Journals Inc.