A light-up G-quadruplex nanostring for label-free and selective detection of miRNA via duplex-specific nuclease mediated tandem rolling circle amplification

MicroRNA (miRNA) has emerged as a promising genetic marker for cancer diagnosis and therapy because its expression level is closely related to the progression of malignant diseases. Herein, a label-free and selective fluorescence platform was proposed for miRNA based on light-up G-quadruplex nanostring via duplex-specific nuclease (DSN) mediated tandem rolling circle amplification (RCA). First, a long DNA generated from upstream RCA was designed with the antisense sequences for miR-21 and downstream RCA primer. Upon recognizing miR-21, the resulting DNA RNA permitted DSN digestion and triggered downstream two-way RCA. and generation of abundant G-quadruplex nanostring binding with ZnPPIX for label-free fluorescent responses. In our strategy, the strong preference of DSN for perfectly matched DNA/RNA ensures its excellent selectivity. The developed method generated wide linear response with LOD of 1.019 fM. Additionally, the miR-21 levels in cell extracts have been evaluated, revealing the utility of this tool for biomedical research and clinical diagnosis. (C) 2020 Elsevier Inc. All rights reserved.

Automated summary

A label-free and selective fluorescence platform for miRNA detection was developed using G-quadruplex nanostring and DSN-mediated RCA. The method showed excellent selectivity for perfectly matched DNA/RNA, with a detection limit of 1.019 fM for miR-21. This tool has potential applications in evaluating miR-21 levels in cell extracts for biomedical research and clinical diagnosis.