Exonuclease III-powered DNA Walking Machine for Label-free and Ultrasensitive Electrochemical Sensing of Antibiotic

Herein, an Exonuclease III (Exo III)-powered DNA walking machine has been integrated into electrochemical sensing for label-free and highly sensitive detection of antibiotics. The electrochemical sensing interface is prepared by co-assembly of hemin aptamer-containing DNA track (DT) and DNA walker (DW)-locking probe (LP) duplex on gold electrode surface. The specific target and aptamer interaction triggers Exo III-assisted cyclic amplification, releasing the unlocking probe (UP) that is used for the deblocking of DW. Then the autonomous moving of DW on electrode surface is propelled via Exo III digestion of hybridized DT. The consumption of DT induces the formation of G-quadruplex/hemin DNAzyme that can catalyze the reduction of H 2 O 2 , thus a significantly strong current signal is released. Using ampicillin as a model analyte, the proposed biosensor achieves a detection limit of 0.76 pM within a linear detection range from 1 pM to 10 nM. Additionally, this biosensing strategy can be readily extended for the detection of wide variety of analytes by using corresponding target recognition probes. Therefore, the proposed strategy indeed creates a highly sensitive and label-free electrochemical sensing platform for molecular diagnosis applications.