Solid-State Nanopore Single-Molecule Sensing of DNAzyme Cleavage Reaction Assisted with Nucleic Acid Nanostructure

The detection and investigation of biomolecules at a single-molecule level is important for improving diagnosis in biomedicine. Solid-state nanopores are a unique tool that have the potential to accomplish this task because they are label-free and require only low sample consumption. However, the event-readouts of current small polymer molecules are still limited because of its relatively large size and low signal-to-noise ratios. Here, we present a rapid sensing approach for the detection of GR-5 DNAzyme cleaving specific substrate reactions using relatively larger size silicon nitride nanopores by introducing a type of nucleic acid nanostructure (DNA tetrahedron) as a carrier. The proposed method is convenient and sensitive enough to detect the cleavage reactions by identifying translocation events before and after reactions with nanomolar concentrations of the target sample. Furthermore, this assay was also carried out by using larger size nanopores (60 nm diameter) to achieve the DNAzyme cleavage sensing with the same sample concentration. This approach can improve event detectability of other smaller molecules' translocation, which opens up a wide range of applications for analytes detection by incorporating solid-state nanopores. Nucleic acid nanostructure-assisted nanopore sensing can promote the development of single-molecule studies.