Enzymatically Amplified Surface Plasmon Resonance Imaging Method Using RNase H and RNA Microarrays for the Ultrasensitive Detection of Nucleic Acids

摘要

A novel surface enzymatic amplification method that utilizes RNA microarrays in conjunction with the enzyme RNase H is developed for the ultrasensitve detection and analysis of target DNA molecules.The enzyme RNase H is shown to selectively and repeatedly destroy RNA from RNA-DNA heteroduplexes on gold surfaces;when used in conjunction with the label-free technique of surface plasmon resonance imaging,multiple DNA targets can be detected at a concentration of 10 fM on a single chip.In addition,this method is utilized for the sequence-specific detection of the TSPY gene in both purified and unpurified PCR products.Finally,in a series of kinetics measurements,the initial rate of hydrolysis is shown to depend directly on the surface concentration of DNA-RNA heteroduplexes.