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Microbial Genomics logo Microbial Genomics

Volume 10, Issue 6

How low can you go? Short-read polishing of Oxford Nanopore bacterial genome assemblies Open Access

Abstract

It is now possible to assemble near-perfect bacterial genomes using Oxford Nanopore Technologies (ONT) long reads, but short-read polishing is usually required for perfection. However, the effect of short-read depth on polishing performance is not well understood. Here, we introduce Pypolca (with default and careful parameters) and Polypolish v0.6.0 (with a new careful parameter). We then show that: (1) all polishers other than Pypolca-careful, Polypolish-default and Polypolish-careful commonly introduce false-positive errors at low read depth; (2) most of the benefit of short-read polishing occurs by 25× depth; (3) Polypolish-careful almost never introduces false-positive errors at any depth; and (4) Pypolca-careful is the single most effective polisher. Overall, we recommend the following polishing strategies: Polypolish-careful alone when depth is very low (<5×), Polypolish-careful and Pypolca-careful when depth is low (5–25×), and Polypolish-default and Pypolca-careful when depth is sufficient (>25×).

  • Received:
  • Accepted:
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Keyword(s): assembly polishing , bacterial genome assembly , genome polishing and Oxford nanopore sequencing
Funding
This study was supported by the:
  • National Health and Medical Research Council (Award GNT1194325)
    • Principle Award Recipient: TimothyP. Stinear
  • Garnett Passe and Rodney Williams Memorial Foundation
    • Principle Award Recipient: SarahVreugde
  • National Institutes of Health (Award RC2DK116713)
    • Principle Award Recipient: RobertA. Edwards
© 2024 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
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2024-06-04
2025-06-24
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/content/journal/mgen/10.1099/mgen.0.001254
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How low can you go? Short-read polishing of Oxford Nanopore bacterial genome assemblies
M Gen 10, 001254 (2024); https://doi.org/10.1099/mgen.0.001254
/content/journal/mgen/10.1099/mgen.0.001254
/content/journal/mgen/10.1099/mgen.0.001254
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