A mismatch PCR-RFLP primer design for SNP genotyping using genetic algorithm

Designing a feasible primer pair is an important work before performing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) for single nucleotide polymorphism (SNP) genotyping. However, in many cases, no restriction enzymes are available to discriminate the target SNP, thus rendering the primer design useless. We propose a method that uses a genetic algorithm (GA) to search for optimal mutagenic PCR-RFLP primers and employ the core of SNP-RFLPing to reliably mine available restriction enzymes. The in silico simulation of the proposed method in the SNPs of the SLC6A4 gene showed that it is capable of designing stable mismatch PCR-RFLP primers which fit the common primer constraints and that it can provide available restriction enzymes.