Improvements in allelic discrimination of microsatellite markers using denaturing polyacrylamide gel electrophoresis

Abstract

Poor resolution, retarded progress of DNA through gels, and variable sizing of DNA fragments between and within gels hinder accurate genotyping of some simple sequence length polymorphism (SSLP) markers with the Perkin Elmer Applied Biosystems 377 Sequenator. These problems are similar to renaturation related problems observed in DNA sequencing gels. PCR products especially susceptible to these problems are shown to have higher melting temperatures (T(m)) than others. Gels containing increased concentrations of denaturants allow greater accuracy in allelic discrimination. This is especially beneficial where quantification is necessary.