Development of a multiplex PCR assay detecting 52 autosomal SNPs

Abstract

An efficient method that can be used to simultaneously amplify a set of genetic loci across the genome with high reliability can provide a valuable tool for single nucleotide polymorphism (SNP) forensic genotyping. A crucial element is the number of individual biochemical reactions that must be performed. The SNPforID consortium (www.snpforid.org) was established in 2003 with the principal goal of developing a SNP-based system of DNA analysis that would have comparable discrimination power and ease of use to those of existing short tandem repeat (STR) based techniques. Here, we describe a strategy for amplifying 52 genomic DNA fragments, each containing one SNP, in a single tube, and accurately genotyping the PCR product mixture using two single base extension reactions. This multiplex approach reduces the cost of SNP genotyping and requires as little as 0.5 ng of genomic DNA to detect 52 SNPs. We used a multiple injection approach for DNA sequencers that can effectively detect all the SNPs amplified in a single electrophoretic run. We present SNP data for 700 unrelated individuals from 9 populations.