Synergetic dual-toehold mediated controllable transcription amplification for detecting lung cancer-related circulating miRNAs in blood

Abstract

Circulating microRNAs (miRNAs) in blood are recognized as key biomarkers of cancers. Herein, taking advantage of the dynamic DNA nanotechnology, we report an assay to detect lung cancer-related circulating miRNAs in blood based on a synergetic dual-toehold mediated controllable transcription amplification (SMART). Unlike traditional single-toehold overhanging system with limited design flexibility, the synergetic dual-toehold exchange was employed to not only inhibit background signal, but also facilitate the downstream introducing of target-responsive isothermal amplification. This feature allows the SMART platform combines synergetic dual-toehold mediated strand displacement with transcription amplification. The SMART, acting as an effective signal transducer and amplifier, is thus able to ultrasensitively output the let-7a recognition event via significantly accumulating Broccoli RNA aptamers responsible for the binding with DFHBI-1 T. Moreover, the Broccoli/DFHBI-1 T reporter increases the signal-to-background ratio compared to that using non-specific nucleic acid dyes and eliminates the use of chemically labeled oligonucleotide probes. The experimental demonstration shows the SMART method realized the detection of let-7a lower into 10 fM, identification of let-7a from homologous and heterologous analogues, and profiling let-7a from clinical blood samples. This work improves the rational design of dynamic DNA devices-based signal isothermal in the fields of disease diagnosis and biomedical study.