High-Resolution Detection of PCR Products from a Microsatellite Marker Using a Nonradioisotopic Technique

Abstract

We report a safe, rapid, and economical method for polymerase chain reaction (PCR)-based genotype analysis using a microsatellite marker specific for the human chromosome 18 locus, D18S53. This method does not involve radioisotopes and makes use of ethidium bromide fluorescence to detect PCR products. Our method enables direct analysis and easy detection of PCR products on nondenaturing polyacrylamide gels. The genotyping using this method can be scaled up to 100 samples at one time by adding a step of “double loading” of samples in a single sequencing size gel. We could resolve PCR products and DNA fragments, differing in size by only 2 bp, in the range of 150–200 bp by a 7% nondenaturing polyacrylamide gel. This technique can be applied for population-based genomic screening and linkage analysis.