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Whole genome sequence and manual annotation of Clostridium autoethanogenum, an industrially relevant bacterium

Humphreys, Christopher M.; McLean, Samantha; Schatschneider, Sarah; Millat, Thomas; Henstra, Anne M.; Annan, Florence J.; Breitkopf, Ronja; Pander, Bart; Piatek, Pawel; Rowe, Peter; Wichlacz, Alexander T.; Woods, Craig; Norman, Rupert; Blom, Jochen; Goesman, Alexander; Hodgman, Charlie; Barrett, David; Thomas, Neil R.; Winzer, Klaus; Minton, Nigel P.

Authors

Samantha McLean

Sarah Schatschneider

Anne M. Henstra

Florence J. Annan

Ronja Breitkopf

Bart Pander

Pawel Piatek

Peter Rowe

Alexander T. Wichlacz

Craig Woods

Rupert Norman

Jochen Blom

Alexander Goesman

Charlie Hodgman

David Barrett

NEIL THOMAS neil.thomas@nottingham.ac.uk
Professor of Medicinal and Biological Chemistry

NIGEL MINTON nigel.minton@nottingham.ac.uk
Professor of Appliedmolecular Microbiology



Abstract

© 2015 Humphreys et al. Background: Clostridium autoethanogenum is an acetogenic bacterium capable of producing high value commodity chemicals and biofuels from the C1 gases present in synthesis gas. This common industrial waste gas can act as the sole energy and carbon source for the bacterium that converts the low value gaseous components into cellular building blocks and industrially relevant products via the action of the reductive acetyl-CoA (Wood-Ljungdahl) pathway. Current research efforts are focused on the enhancement and extension of product formation in this organism via synthetic biology approaches. However, crucial to metabolic modelling and directed pathway engineering is a reliable and comprehensively annotated genome sequence. Results: We performed next generation sequencing using Illumina MiSeq technology on the DSM10061 strain of Clostridium autoethanogenum and observed 243 single nucleotide discrepancies when compared to the published finished sequence (NCBI: GCA_000484505.1), with 59.1 % present in coding regions. These variations were confirmed by Sanger sequencing and subsequent analysis suggested that the discrepancies were sequencing errors in the published genome not true single nucleotide polymorphisms. This was corroborated by the observation that over 90 % occurred within homopolymer regions of greater than 4 nucleotides in length. It was also observed that many genes containing these sequencing errors were annotated in the published closed genome as encoding proteins containing frameshift mutations (18 instances) or were annotated despite the coding frame containing stop codons, which if genuine, would severely hinder the organism's ability to survive. Furthermore, we have completed a comprehensive manual curation to reduce errors in the annotation that occur through serial use of automated annotation pipelines in related species. As a result, different functions were assigned to gene products or previous functional annotations rejected because of missing evidence in various occasions. Conclusions: We present a revised manually curated full genome sequence for Clostridium autoethanogenum DSM10061, which provides reliable information for genome-scale models that rely heavily on the accuracy of annotation, and represents an important step towards the manipulation and metabolic modelling of this industrially relevant acetogen.

Citation

Humphreys, C. M., McLean, S., Schatschneider, S., Millat, T., Henstra, A. M., Annan, F. J., …Minton, N. P. (2015). Whole genome sequence and manual annotation of Clostridium autoethanogenum, an industrially relevant bacterium. BMC Genomics, 16(1), https://doi.org/10.1186/s12864-015-2287-5

Journal Article Type Article
Acceptance Date Dec 10, 2015
Online Publication Date Dec 21, 2015
Publication Date Dec 21, 2015
Deposit Date Jan 14, 2016
Publicly Available Date Jan 14, 2016
Journal BMC Genomics
Electronic ISSN 1471-2164
Publisher Springer Verlag
Peer Reviewed Peer Reviewed
Volume 16
Issue 1
Article Number 1085
DOI https://doi.org/10.1186/s12864-015-2287-5
Keywords Clostridium autoethanogenum, Next generation sequencing, Acetogen, Manual annotation, Synthesis gas fermentation
Public URL http://eprints.nottingham.ac.uk/id/eprint/31246
Publisher URL http://bmcgenomics.biomedcentral.com/articles/10.1186/s12864-015-2287-5
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0

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