title: DNA stretching and optimization of nucleobase recognition in enzymatic nanopore sequencing.
abstract: In nanopore sequencing, where single DNA strands are electrophoretically translocated through a nanopore and the resulting ionic signal is used to identify the four DNA bases, an enzyme has been used to ratchet the nucleic acid stepwise through the pore at a controlled speed. In this work, we investigated the ability of alpha-hemolysin nanopores to distinguish the four DNA bases under conditions that are compatible with the activity of DNA-handling enzymes. Our findings suggest that in immobilized strands, the applied potential exerts a force on DNA (∼10 pN at +160 mV) that increases the distance between nucleobases by about 2.2 Å V(-1). The four nucleobases can be resolved over wide ranges of applied potentials (from +60 to +220 mV in 1 m KCl) and ionic strengths (from 200 mM KCl to 1 M KCl at +160 mV) and nucleobase recognition can be improved when the ionic strength on the side of the DNA-handling enzyme is low, while the ionic strength on the opposite side is high.
authors: University of Oxford, Chemistry Research Laboratory, Oxford, UK.
authors: David
authors: D
authors: Stoddart
authors: Lorenzo
authors: L
authors: Franceschini
authors: Andrew
authors: A
authors: Heron
authors: Hagan
authors: H
authors: Bayley
authors: Giovanni
authors: G
authors: Maglia
mesh_terms: D004247:DNA / Q000737:chemistry*; D004560:Electricity; D006863:Hydrogen-Ion Concentration; D007477:Ions; D058608:Nanopores; D017422:Sequence Analysis, DNA / Q000379:methods*; D013314:Stress, Mechanical
chemical_list: D007477:Ions; D004247:DNA
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