Establishment and application of a multiple allele-specific PCR method based on a novel high-fidelity Taq polymerase
This technology was established on the basis of three selected SNP loci for preliminary testing on mice with partially re-placed fragments on chromosome 8,including specificity testing of high-fidelity Taq enzymes and evaluation of the effect of primer con-centration on genotyping results as well as sensitivity assessment.It was used to detect nine SNPs in different wild-type chromosome 1 substitution mouse samples.The study showed that the detection results without introducing mutations into primers were consistent with sequencing results.The dilution experiments indicated that the optimal concentration was at least 0.003-0.006 μmol/L,while sensitivity detection revealed that the minimal concentration detection limit could reach below 0.07 ng/μL.Finally,five standard sam-ples'genotypes representing combinations of nine SNPs were detected.The overall process for applying this highly specific Taq enzyme in multiple allele-specific PCR experiments was eventually validated and established.This article offered a low-cost solution with high specificity and the ability to detect multiple SNP loci,which preserved the reference value for the development of multiple allele-specif-ic PCR technology.
万方数据
维普
