Discriminating Bases by Stretching Double-Stranded DNA in a Nanopore

Abstract

We report a new method for trapping a single molecule of double-stranded DNA (dsDNA) in a solid-state nanopore in SiNx membrane and describe the prospects for sequencing it. It is possible to trap a single dsDNA molecule in a nanopore <3nm in diameter by first applying a voltage larger than this threshold and forcing the molecule to translocate through the pore. If the electric field is then rapidly switched to a value below threshold, the DNA becomes trapped for seconds in the pore, compared to a sub-millisecond translocation if the field is maintained above threshold. Moreover, if the duration in the trap is commensurate with the bandwidth we can discriminate distinct signatures of C-G and A-T base-pairs by simply measuring the pore current. Molecular dynamics simulations of these experiments reveal that, when trapped, the dsDNA is stretched in the pore in a specific tilted orientation, depending on the orientation of the leading nucleotides, while the B-form canonical structure is preserved outside the pore. Finally, we show using streptavidin bound biotinylated dsDNA (Fig. 1a) that it's possible to discriminate stretched basepairs in the trapped configuration (Fig. 1b,c) between A-T and C-G.View Large Image | View Hi-Res Image | Download PowerPoint Slide