Single-Molecule Signatures for Characterization and Regulation of Nucleic Acids Unzipping in a Nanopore

Abstract

The nanopore has been developed as a molecule force microscope to explore the unzipping of nucleic acids. The double-stranded DNA (dsDNA) such as hairpin with an overhang trapped in the nanopore can generate long current blocks. By measuring the voltage-dependent block duration, the unzipping kinetics as well as the force and energy involved in the double strands hybridization can be characterized. However, few studies have presented convincing characteristic current patterns for unzipping occurrence. In this report, we uncovered such a signature current pattern that can electrically track the entire unzipping process, from the time course of unzipping to the motion pathway of unzipped single-stranded DNA. With the signature signals, the release of DNA without unzipping as well as DNA trapping directionality can also be distinguished. Quantitative analysis of signature signals showed that the overhang of a DNA is more favorable to the nanopore compared to the blunt end. Therefore the overhang is not only an unzipping driver, but also a controller of DNA trapping orientation, regulating the unzipping in a programmable manner. Discrimination of unzipping signatures not only gives precise insight into the unzipping mechanism, but more importantly in biosensors, the unzipping signature can act as a single-molecule marker that ensures both selectivity and sensitivity.