Enzyme-free colorimetric method to detect ribonuclease H activity based on toehold-mediated strand displacement reaction

Abstract

We herein developed a novel enzyme-free and label-free colorimetric method to determine ribonuclease H (RNase H) activity utilizing target-induced toehold-mediated strand displacement (TMSD) reaction. In principle, this strategy employs a detection probe, which is an RNA/DNA hybrid duplex probe, to determine RNase H activity. RNase H hydrolyzes the RNA strand in the detection probe and liberates a catalyst strand (CS), which initiates the TMSD reaction. Released DNA strand liberates a G-quadruplex strand (GS), which is initially caged by a blocker strand, and repeatedly participates in TMSD reaction with the aid of fuel strand (FS). As a result, an intense colorimetric signal is produced from ABTS oxidized by the peroxidase mimicking activity of G-quadruplex. With this method, we sensitively determined the RNase H activity down to 0.27 U/ml with high selectivity against other nucleases and restriction enzyme.