Direct single-nucleotide polymorphism genotyping from whole blood without DNA extraction

A subset of single-nucleotide polymorphisms (SNPs) are associated with various phenotypes, including diseases, coat colour, and athletic performance, and are widely used in medical and veterinary fields. In this study, we developed a novel method for direct SNP genotyping from whole blood without DNA extraction. Thoroughbred blood samples were diluted 100-fold with Milli-Q water and analysed using real-time polymerase chain reaction (PCR) with hydrolysis probes. Specificity and sensitivity were improved by increasing the annealing temperature and number of PCR cycles. Genotyping results for SNPs in <i>MSTN</i> and <i>LCORL</i> showed complete concordance with conventional real-time PCR using the extracted DNA. This method is simple, low-cost, highly versatile, and applicable to other genetic targets, such as <i>CDH13</i> and <i>MAOA</i>.