Concourse of Primer Design

Knowledge of single nucleotide polymorphisms (SNPs) constitutes essential information in studies in personalized medicine. Many kinds of software are developed for SNP genotyping. However, many of them require costly reagents and investments. Here, we introduce a web-based software, "Prim-SNPing", which provides three kinds of primer design for cost-effective SNP genotyping, including natural primer design (PD), mutagenic PD and CTPP (PCR with confronting two-pair primers) PD. The natural and mutagenic PDs are restriction enzyme-required for restriction fragment of length polymorphism (RFLP), while the CTPP PD is restriction enzyme-free after PCR. The PCR efficiency for the designed primers is enhanced by BLAST and quality control avoiding secondary structures. For novelty, the length pattern of RFLP using natural PD is user-adjustable and the provided RFLP enzymes are confirmed to be absent within the generated PCR product. In CTPP, it eliminates the need for a separate digestion step in RFLP, and is thereby faster and cheaper to perform. The output for Prim-SNPing includes the primer list, Tm value, GC% and amplicon size with or without enzyme digestion. NCBI dbSNP rs IDs and/or free form of SNP sequence formats from the human, mouse and rat genomes are acceptable for Prim-SNPing. Therefore, the Prim-SNPing provides an interactive, user-friendly and cost-effective primer design for SNP genotyping commonly used in regulatory laboratories.