A self-powered 3D DNA walking machine triggered by triple-helix molecular switch for microRNA imagining in living cells

Herein, a novel self-powered 3D DNA walking machine triggered by triple-helix molecular switch (THMS) has been proposed for microRNA-21 (miRNA-21) imaging in living cells. Impressively, the 3D DNA walking machine was constructed with swing arm and THMS co-functionalized gold nanoparticles, THMS could precisely customize as a multifunctional structure with variable configuration. Once the 3D DNA walking machine was delivered into living cells, THMS was promptly disassembled due to the formation of DNA duplex (miRNA-21/TAMRA-DNA2), thus the 3D DNA walking machine was immediately triggered on AuNPs. Meanwhile, FAM-DNA1 quickly self-hybridized into hairpin and exposed the Mg2+ ribonucleoside adenosine (rA) recognition site in the loop region. Sequentially, swing arm hybridized with hairpin FAM-DNA1 and cleaved it with the assistance of Mg2+. Finally, numerous FAM-labeled segments were released from AuNPs and further paired with the spare part of TAMRA-DNA2 in the miRNA-21/TAMRA-DNA2 duplex, resulting in the two fluorescent dyes approaching each other, which induced the efficient FRET from FAM to TAMRA, thereby turned the system into “ON” state for signal amplification. It is noteworthy that the ingenious design of the self-powered 3D DNA walking machine avoids the external addition of fuel DNA strands or protein enzymes, which enables the autonomous motion of the 3D DNA walking machine in complex cellular microenvironment, greatly simplifies the experimental steps and improves the efficiency of the walking machine. Moreover, this assay will provide a novel signal amplification strategy for evaluating intracellular miRNA levels.