A split G-quadruplex-based DNA nano-tweezers structure as a signal-transducing molecule for the homogeneous detection of specific nucleic acids.

A portable method of specific nucleic acid detection would be very useful for monitoring public health in a variety of settings for point-of-care and point-of-need testing. However, conventional methods for the detection of nucleic acids are not ideal for use in the field, as they require skilled operators and complex equipment. Here, we constructed a method for specific nucleic acid detection using a split G-quadruplex (Gq) structure that can recognize target nucleic acids without competitive reactions in a bimolecular reaction and directly produce a detectable signal based on peroxidase activity. We developed a single signal-transducing molecule with a split Gq-based DNA-nano tweezers (NT) structure that self-assembles from three single-stranded DNAs through simple mixing, and detects its target without requiring any washing steps. A model target, a partial norovirus mRNA (NV-RNA), was specifically recognized by the split Gq-based DNA-NT, causing it to undergo a structural change that restored its peroxidase activity. The peroxidase activity was measured by following the oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), which gave a greenish colorimetric response, and was proportional to the NV-RNA concentration. The lower detection limit was 4 nM. Our results demonstrated the feasibility of detecting specific nucleic acids with a split Gq-based DNA-NT structure as a nucleic acid signal-transducing molecule in a homogenous assay format. Also the target recognition sites of split Gq-based DNA-NT can easily be designed without delicate optimization of tweezers structure. Thus a split Gq-based DNA-NT technique is readily applicable to a basic platform for the development of a portable device.