Controlling DNA Tug‐of‐War in a Dual Nanopore Device

Abstract

Methods for reducing and directly controlling the speed of DNA through a nanopore are needed to enhance sensing performance for direct strand sequencing and detection/mapping of sequence‐specific features. A method is created for reducing and controlling the speed of DNA that uses two independently controllable nanopores operated with an active control logic. The pores are positioned sufficiently close to permit cocapture of a single DNA by both pores. Once cocapture occurs, control logic turns on constant competing voltages at the pores leading to a “tug‐of‐war” whereby opposing forces are applied to regions of the molecules threading through the pores. These forces exert both conformational and speed control over the cocaptured molecule, removing folds and reducing the translocation rate. When the voltages are tuned so that the electrophoretic force applied to both pores comes into balance, the life time of the tug‐of‐war state is limited purely by diffusive sliding of the DNA between the pores. A tug‐of‐war state is produced on 76.8% of molecules that are captured with a maximum two‐order of magnitude increase in average pore translocation time relative to the average time for single‐pore translocation. Moreover, the translocation slow‐down is quantified as a function of voltage tuning and it is shown that the slow‐down is well described by a first passage analysis for a 1D subdiffusive process. The ionic current of each nanopore provides an independent sensor that synchronously measures a different region of the same molecule, enabling sequential detection of physical labels, such as monostreptavidin tags. With advances in devices and control logic, future dual‐pore applications include genome mapping and enzyme‐free sequencing.

Flossing DNA in a Dual Nanopore Device

“…The control logic was run in real‐time (MHz clock rate) on a field programmable gate array (FPGA). We modified our previously designed tug‐of‐war control process to permit loading reagent in the common fluidic chamber above the two nanopores and to screen out short‐fragments (Section S2 and Figure S1, Supporting Information). Once co‐capture is achieved, the competing voltage forces at V 1 and V 2 lead to a tug‐of‐war and reduce the DNA speed during sensing.…”

“…While prior work leveraged control logic for automated recapture and sensing of molecules with single nanopores, our approach is distinctive in a few ways. First, our device employs a dual‐pore architecture . By having the dual pores sufficiently close, our prior work shows the DNA can be captured simultaneously by both pores and exist in a “tug‐of‐war” state where competing electrophoretic voltage forces are applied at the pores .…”

“…First, our device employs a dual‐pore architecture . By having the dual pores sufficiently close, our prior work shows the DNA can be captured simultaneously by both pores and exist in a “tug‐of‐war” state where competing electrophoretic voltage forces are applied at the pores . During tug‐of‐war, the molecule's orientation and identity are maintained, the molecule speed is regulated to facilitate tag sensing, and the likelihood of the molecule finding a linearized conformation through the pore is increased to 70% compared to 30% with single‐pore data.…”

Controlling DNA Translocation Through Solid-state Nanopores

“…The doublenanopore system opens up a new path to the mechanical trapping of DNA in solid-state nanopores, and it is a promising technique to measure a wide range of biomolecules with the advantages of being label-free, and having a high signal-to-noise ratio and low cost. It can efficiently confine and trap the DNA molecules to slow down DNA translocation and can also be used to study the physics of this nanoscale tug-of-war on DNA [41].…”

Single Molecule Protein Sequencing Based on the Superspecificity of tRNA Synthetases

“…A range of methods to slow down the polymer in its passage through the nanopore have been reported, an early review can be found in [32] (but also see [33]). More recent work can be found in [34][35][36][37][38][39][40]. The primary aim in all of the above slowdown methods is to allow discrimination of bases in DNA and residues in proteins (and, in one case, unfolding of the protein as well [34]).…”