Exponential Strand-Displacement Amplification for\nDetection of MicroRNAs

Abstract

MicroRNAs\n(miRNAs) are promising targets for disease diagnosis. However, miRNA\ndetection requires rapid, sensitive, and selective detection to be\neffective as a diagnostic tool. Herein, a miRNA-initiated exponential\nstrand-displacement amplification (SDA) assay was reported. With the\nKlenow fragment, nicking enzyme <i>Nt.AlwI</i>, and two\nprimers, the miRNA target can trigger two cycles of nicking, polymerization,\nand displacement reactions. These reaction cycles amplified the target\nmiRNA exponentially and generated dsDNAs detectable with SYBR Green\nI in real-time PCR. As low as 16 zmol of the target miRNA was detected\nby this one-pot assay within 90 min, and the dynamic range spanned\nover 9 orders of magnitude. Negligible impact from the complex biological\nmatrix was observed on the amplification reaction, indicating the\nassay’s capability to directly detect miRNAs in biofluids.