Affordable Access

deepdyve-link
Publisher Website

Automated forward and reverse ratcheting of DNA in a nanopore at 5-Å precision.

Authors
  • Gm, Cherf
  • Kr, Lieberman
  • H, Rashid
  • Ce, Lam
  • Kevin Karplus
  • M, Akeson
Type
Published Article
Journal
Nature Biotechnology
Publisher
Springer Nature
Volume
30
Issue
4
Pages
344–348
Identifiers
DOI: 10.1038/nbt.2147
Source
UCSC Bioinformatics biomedical-ucsc
License
Unknown

Abstract

An emerging DNA sequencing technique uses protein or solid-state pores to analyze individual strands as they are driven in single-file order past a nanoscale sensor. However, uncontrolled electrophoresis of DNA through these nanopores is too fast for accurate base reads. Here, we describe forward and reverse ratcheting of DNA templates through the α-hemolysin nanopore controlled by phi29 DNA polymerase without the need for active voltage control. DNA strands were ratcheted through the pore at median rates of 2.5-40 nucleotides per second and were examined at one nucleotide spatial precision in real time. Up to 500 molecules were processed at ∼130 molecules per hour through one pore. The probability of a registry error (an insertion or deletion) at individual positions during one pass along the template strand ranged from 10% to 24.5% without optimization. This strategy facilitates multiple reads of individual strands and is transferable to other nanopore devices for implementation of DNA sequence analysis.

Report this publication

Statistics

Seen <100 times