Controllable Linear DNA Nanomotor Enables Ultrasensitive Detection of MicroRNA

Abstract

The catalytic hairpin assembly amplification strategy, owing to its designability and high specificity, has been widely used for highly sensitive detection of low-abundance miRNAs. However, uncontrollable valency and distance between the hairpin probes result in low reaction rates and efficiency, often necessitating extended incubation times, thereby limiting their practical applications. In this study, we present a controllable linear DNA nanomotor (CLDN) composed of a pair of interval hybridization-modified hairpin DNA probes (H1 and H2) for the highly sensitive and selective detection of miRNAs. By regulating both reaction time and sequence interval, the length of the DNA scaffold generated by rolling circle amplification can be controlled, thus modulating the assembly valency and distance between the hairpin DNA probes. When target miRNAs sequentially initiate the interval hybridization of H1 and H2, the entire DNA nanomotor is rapidly activated through the simultaneous opening of all self-quenching hairpins (H1). Owing to the acceleration from the confined space and domino-like effect, the signal-to-noise ratio is enhanced by approximately 6.73-fold in comparison to conventional DNA cascade reaction. Additionally, the CLDN demonstrates high selectivity in distinguishing between homogeneous miRNA sequences with a single-base difference. Our CLDNs hold great potential for quantitatively analyzing multiple miRNAs in clinical diagnostics.

Keywords

DNA structures / Catalytic hairpin assembly / Rolling circle amplification / Confined space / Fluorescence / Biosensing

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2025 SIOC, CAS, Shanghai, & WILEY-VCH GmbH