RCA-HCR cascade mediated-linear multi-shear site DNA walker (LMDW) to construct an electrochemical biosensor for ultrasensitive detection of Tumor markers.

Herein, through the cascade integration of the RCA and HCR reactions, an innovative linear multi-shear site DNA walker (LMDW) with high walking efficiency was developed to fabricate an electrochemical biosensing platform for detecting microRNA-21 (miRNA-21) associated with breast cancer. Impressively, compared with conventional single-shear site DNA walkers with poor walking efficiency, the LMDW obtained from the RCA-confined HCR reaction can accommodate a considerable number of functional domains, increasing the shear efficiency to the hairpin probe with the beacon material, thereby consequently enhancing the reaction rate and sensitivity of the biosensor. Following validation, the probe acquisition time down to just 20 min, far less than traditional single-shear site DNA walkers and the detection limit was 0.72 aM, which was much lower than that reported in the literature. In addition, the electrochemical biosensor based on LMDW exhibited unbelievable reproducibility, stability and selectivity, and demonstrated effective application in the detection of miRNA-21 in MCF-7 and HeLa cell lysates, displaying high expression in the former and low expression in the latter, demonstrating excellent potential for application, promoting the research on the intrinsic performance of nucleic acid signal cascade amplification, and providing a new perspective for miRNA detection in biological samples and eventual clinical disease diagnosis.