Highly accurate analysis of heterozygous loci bysingle cell PCR

Abstract

Single cell PCR is a powerful method for determining the genetic properties of individual cells. In the instance of heterozygous loci, however, preferential amplification of one allele can lead to allele drop out (ADO) of the other allele. Fortunately ADO does not occur in all amplifications, and is usually random when it does occur, with both alleles being equally susceptible to drop out. Therefore pooling of results from multiple independently amplified cells should greatly improve the analysis of diallelic loci, and the misdiagnosis rate of diallelic loci should decrease exponentially with the number of cells analysed. We have shown that this is true and that multiplex PCR allows for the simultaneous identification of a cell in a mixture of cells using microsatellite loci known to be informative, and accurate genotyping at other loci. This approach has practical applications to non-invasive prenatal diagnosis where small numbers of fetal cells in the presence of maternal cells must be both identified and genotyped at loci involved in genetic disease.