An artificial-restriction fragment length polymorphism (A-RFLP) method for genotyping intronic SNP rs7587633 C/T of ATG16L1.

Abstract

SNPs could either cause a disorder or directly alter the efficacy of a particular treatment and act as biological markers. The SNP rs7587633 C/T present in the intronic region of the ATG16L1 gene has been studied for its role in psoriasis vulgaris and Palmoplantar pustulosis. To genotype rs7587633 C/T using PCR-RFLP no restriction site is present for any of the restriction enzymes at the SNP position. To develop an artificial-RFLP method for genotyping rs7587633 C/T, the forward primer was designed in such a way that it resulted in the creation of an EcoRI restriction site in the amplified product which could further be digested with EcoRI to find the genotype of the individual. The newly developed A-RFLP method was applied to genotype the SNP rs7587633 C/T in DNA samples of 100 healthy control individuals. The allelic and genotypic frequencies of the SNPs were 0.80(C), 0.20(T) and 65%(CC), 31%(CT) and 4%(TT), respectively. In conclusion, we developed an A-RFLP method to genotype the SNP rs7587633 C/T which is not present in any of the natural restriction sites and this method could be applied to genotype this SNP in various populations/diseases to find its role.