Substantial Slowing of Electrophoretic Translocation of DNA through a Nanopore Using Coherent Multiple Entropic Traps

Abstract

One of the major challenges in the technology of sequencing DNA using single-molecule electrophoresis through a nanopore is to control the translocation of the macromolecule across the pore in order to allow sufficient time for accurate sequence reading at limited recording bandwidths. If the translocation speed is too fast, the signatures of the bases passing through the sensing region of the nanopore overlap in time, presenting difficulties in accurately identifying the bases in a sequential manner. Even though several strategies, such as enzyme ratcheting, have been implemented to reduce the translocation speed, the challenge to achieve a substantial reduction in the translocation speed continues to be of paramount significance. Toward achieving this goal, we have fabricated a nonenzymatic hybrid device that can reduce the translocation speed of long DNAs by more than 2 orders of magnitude, in comparison with the current status of the art. This device is made of a tetra-PEG hydrogel that is chemically anchored to the donor side of a solid-state nanopore. The idea behind this device is based on the recent discovery of the topologically frustrated dynamical state of confined polymers, whereby the front hydrogel matter of the hybrid device provides multiple entropic traps for a single DNA molecule holding it back against the electrophoretic driving force that pulls the DNA through the solid-state nanopore portion of the device. As a demonstration of slowing DNA translocation by a factor of about 500, we find the average translocation time realized in the present hybrid device for 3 kbp DNA as 23.4 ms, whereas the corresponding time for the bare solid-state nanopore under otherwise identical conditions is 0.047 ms. Our measurements on 1 kbp DNA and λ-DNA show that such a slowing down of DNA translocation with our hybrid device is general. An additional feature of our hybrid device is its incorporation of all features of the conventional gel electrophoresis to separate different DNA sizes in a clump of DNAs and to streamline them in an orderly and slow manner into the nanopore. Our results suggest the high potential of our hydrogel-nanopore hybrid device in further advancing the single-molecule electrophoresis technology to accurately sequence very large biological polymers.

Considerable slowdown of short DNA fragment translocation across a protein nanopore using pH-induced generation of enthalpic traps inside the permeation pathway

“…28 Various strategies were devised to slow down translocation, including the incorporation of motor proteins and enzymes, alterations of experimental conditions, or site-directed mutagenesis. 29–54…”

Advances in Hydrogel-Based Drug Delivery Systems

“…This variability is especially pronounced in hydrogels formed through free-radical polymerization. Achieving a uniform mesh size is possible by using symmetrical tetrahedron-like macromeres of identical size for gelation, leading to hydrogels with a more uniform mesh size [87][88][89]. For smaller drug molecules, release is primarily governed by diffusion, allowing for their free movement within the network.…”

Slip-Coupled Electroosmosis and Electrophoresis Dictate DNA Translocation Speed in Solid-State Nanopores

“…Subsequently, the first experimental measurements of the force acting on the DNA in a solid-state nanopore were reported by Lemay and co-workers , by combining optical tweezers with ionic-current detection. In an attempt to use geometrical asymmetry as a tool to restrain the translocation speed of the DNA in such experimental frameworks within preferential threshold limits, the successful usage of conical nanopores was reported by a sequel of studies. − Other approaches for the same purpose delved into different surface modification methods, such as polyelectrolyte layer (PEL) covering, , hydrophobicity, , field-effect, , surface charge modulation, sequential polymerization, and bonding of synthesized hydrogel to the outer surface of the nanochannel, in an effort to enhance the controllability on DNA migration. However, fundamental analytical insights on the underlying transport mechanisms remained to be elusive to a large extent with a lack of insight into the role of the intrinsic wettability of the DNA molecule unless simplistic assumptions were resorted to.…”