A novel method for SNP detection based on combinative analysis of primer extension and ligation

We described a novel method that combined primer extension and ligation for simultaneously detecting a set of single nucleotide variations. To verify the feasibility of this method, seven synthetic templates containing seven KRAS mutation sites and one template without mutation were detected by four primer extension cycles and one ligation reaction in the present of labeled or unlabeled nucleotides. The primer extension cycles and ligation reaction would give three sets of fluorescent signals. By combining the three set of fluorescent signals, each mutations could be discriminated with great accuracy. The results demonstrated that all the mutation sites were discriminated with great accuracy. This method had the potential in simultaneous discriminating various SNPs in a microarray as long as a set of combinations of labeled/unlabeled nucleotides were designed appropriately. It will provide researcher with a high-throughput, time-saving, and labor-saving diagnosis tool.