Allele-Specific Mutation Genotyping with Mismatches in Primer Design

Genotyping technologies for single nucleotide polymorphisms (SNPs) and other mutation types have evolved to become essential tools in various fields. Although high-throughput genotyping technologies occupy a key position in handling large amounts of SNP data, simple, low-cost, and conventional genotyping technologies remain in demand. Allele-specific (AS) polymerase chain reaction (PCR) and its related improved methods can effectively identify target SNPs and allele types using AS primers that introduce instability through mismatched bases at and around the SNP site. In this chapter, we present what is known from the literature on primer design with mismatches for AS-PCR and describe three cases of mutation detection (SNPs and insertions/deletions) associated with functional genes of crop species, which could be useful to guide future AS-PCR experiments.