Guanine-wire based walking machine for highly sensitive and selective detection of circulating microRNA

Abstract

A novel DNA machine-based fluorescent system is designed for highly sensitive and selective detection of circulating microRNA. It is based on topological assembly of Guanine (G)-tetrads and Mg2+ ion-dependent DNAzyme. With the aid of Mg2+ and K+ ions, G-rich sequences first form parallel G-tetrads and further assemble into aggregated G-wires. G-wire is used as not only the walking track but also the quencher of the fluorophore immobilized on its 5′-end. DNAzyme serves as the walker and is embedded in the stem of the tailored hairpin probe in which a recognition element of target microRNA is designed in the loop region. In the absence of target microRNA, the hairpin probe keeps closed to block the DNAzyme walker. In contrast, the presence of target microRNA results in the open of the hairpin probe by hybridizing with the recognition element and the release of the DNAzyme walker. The released walker binds with and cleaves the fluorescence substrate to free the labeled-fluorophore from the track and then steps toward the neighbored substrate. The cycles of this process continuously turn on the fluorophore for the amplified detection. Using this “signal on” strategy, a detection limit of 57.26 fM is realized with five dynamic ranges and high selectivity. Besides, this detection is featured with null protein enzyme and free of labeled-quencher. Thus our system provides a promising candidate for microRNA detection in clinical diagnosis.