Mismatch cleavage detects pathogenic microorganisms.

Abstract

When a DNA probe hybridizes a DNA target and generates a G/A mismatch in the probe-target DNA heteroduplex, the mismatch enzyme, mutY, will cut the A base at the site of the mismatch. This specific cleavage at the mismatched A on the known probe will reveal the complementary DNA sequences of the targets. This study shows mismatch cleavage assays identify the complementary sequence of cryptic plasmid target in the extract of chlamydia infected cells in culture. In addition, the specific cleavage at a single base permits differentiation of two sequences with one base difference. This was shown in differentiating the subtypes of human immunodeficiency virus (HIV) type 1. The addition of amines in the assays increases the sensitivity by freeing the target for recycling. The combined assay system of high sensitivity and demonstrated specificity allows further evaluation for direct identification of pathogenic microorganisms in patient samples.