Simple Tripedal DNA Walker Prepared by Target-Triggered Catalytic Hairpin Assembly for Ultrasensitive Electrochemiluminescence Detection of MicroRNA

Abstract

In contrast to common DNA walkers, multipedal DNA walkers exhibit larger walking area and faster walking kinetics and provide increased amplification efficiency. Consequently, they have received a considerable amount of attention in biosensing. However, most of them are synthesized by immobilizing multiple DNA walking strands on the surface of Au nanoparticles, which is tedious and time-consuming. Simple preparation of multipedal DNA walkers remains a challenge. Herein, we adopted a simple enzyme-free targettriggered catalytic hairpin assembly (CHA) circuit to synthesize a tripedal DNA walker. By walking on a DNA track-functionalized electrode, a sensitive electrochemiluminescence DNA nanomachine biosensor was constructed for sensing miRNA-21. The DNA walker was powered by toehold-mediated strand displacement; the whole process did not need the assistance of enzymes, thus avoiding tedious procedures and enzyme degradation under unfavorable environmental conditions. Specifically, a superior detection limit of 4 aM and a broad linear range of 10 aM to 1 pM were achieved. This CHA−tripedal DNA walker biosensor was then applied for the detection of miRNA-21 in human serum and showed high selectivity and excellent reproducibility, demonstrating its practical application in bioanalysis. In particular, the Y-shaped tripedal DNA walker comes from the DNA circuit, which makes the approach ideally suited for biosensing of small nucleic acid targets.

Living-Cell MicroRNA Imaging with Self-Assembling Fragments of Fluorescent Protein-Mimic RNA Aptamer

“…MicroRNA-21 (miR-21) was chosen as the target RNA because of its biological roles. 39 It turns out that probe I and II split between the 17th and 18th bases have the maximum fluorescence enhancement after incubation with DFHO and miR-21. The farther the split site from the 17th base, the smaller the fluorescence enhancement of DFHO.…”

“…We first split Corn into different G-quadruplex-related positions (from the 5′-end; the sequence of Corn is shown in Table ), then extended the two split fragments each with a four-base-pair neck and 10-base RNA arms that were complementary to the target RNA (Figures and S2). MicroRNA-21 (miR-21) was chosen as the target RNA because of its biological roles . It turns out that probe I and II split between the 17th and 18th bases have the maximum fluorescence enhancement after incubation with DFHO and miR-21.…”

Toehold-mediated strand displacement coupled with single nanoparticle dark-field microscopy imaging for ultrasensitive biosensing

“…4,5 Owing to its characteristics of low abundance, homology in sequences and easy degradation, much effort has been made to achieve high sensitivity and high selectivity in miRNA sensing. Presently, many powerful analytical methods have been developed for miRNA detection, such as electrochemistry, 6,7 electrochemiluminescence, [8][9][10] fluorescence, 11,12 and the realtime quantitative polymerase chain reaction. 13 As a frequently used isothermal amplification assay in molecular biology, toehold-mediated strand displacement (TMSD) usually has high sensitivity and selectivity for nucleic acid recognition, and can identify single base mutations.…”

A sensitive electrochemical DNA sensor based on reduced graphene oxide modified electrode

“…[17] Among them, the combination of electrochemical sensors and DNA walkers has become a hot topic of current research due to their good programmability, ultra-high sensitivity, and excellent biocompatibility. [13,14,18] DNA walker is a smooth and gradual movement of walker strand along a specific path under the influence Binxiang Xu and Yuping Hu contributed equally to this study. of enzymes, environmental stimuli, strand replacement reactions, enabling molecular computation, synthesis of programs, signal generation, and constant amplification.…”