An Exonuclease III-Powered, On-Particle Stochastic DNA Walker

DNA-based machines have attracted rapidly growing interest owing to their potential in drug delivery, biocomputing, and diagnostic applications. Herein, we report a type of exonuclease III (Exo III)-powered stochastic DNA walker that can autonomously move on a spherical nucleic acid (SNA)-based 3D track. The motion is propelled by unidirectional Exo III digestion of hybridized DNA tracks in a burnt-bridge mechanism. The operation of this Exo III-propelled DNA walker was monitored in real time and at the single-particle resolution using total internal reflection fluorescence microscopy (TIRF). We further interrogated the morphological effect of the 3D track on the nuclease activity, which suggested that the performance of the DNA walker was critically dependent upon the DNA density and the track conformation. Finally, we demonstrated potential bioanalytical applications of this SNA-based stochastic DNA walker by exploiting movement-triggered cascade signal amplification.