title: Use of low-coverage, large-insert, short-read data for rapid and accurate generation of enhanced-quality draft Pseudomonas genome sequences.
abstract: Next-generation genomic technology has both greatly accelerated the pace of genome research as well as increased our reliance on draft genome sequences. While groups such as the Genomics Standards Consortium have made strong efforts to promote genome standards there is a still a general lack of uniformity among published draft genomes, leading to challenges for downstream comparative analyses. This lack of uniformity is a particular problem when using standard draft genomes that frequently have large numbers of low-quality sequencing tracts. Here we present a proposal for an "enhanced-quality draft" genome that identifies at least 95% of the coding sequences, thereby effectively providing a full accounting of the genic component of the genome. Enhanced-quality draft genomes are easily attainable through a combination of small- and large-insert next-generation, paired-end sequencing. We illustrate the generation of an enhanced-quality draft genome by re-sequencing the plant pathogenic bacterium Pseudomonas syringae pv. phaseolicola 1448A (Pph 1448A), which has a published, closed genome sequence of 5.93 Mbp. We use a combination of Illumina paired-end and mate-pair sequencing, and surprisingly find that de novo assemblies with 100x paired-end coverage and mate-pair sequencing with as low as low as 2-5x coverage are substantially better than assemblies based on higher coverage. The rapid and low-cost generation of large numbers of enhanced-quality draft genome sequences will be of particular value for microbial diagnostics and biosecurity, which rely on precise discrimination of potentially dangerous clones from closely related benign strains.
authors.affiliation: Department of Cell and Systems Biology, University of Toronto, Toronto, Ontario, Canada. heath.obrien@utoronto.ca
authors.forename: Heath E
authors.initials: HE
authors.lastname: O'Brien
authors.forename: Yunchen
authors.initials: Y
authors.lastname: Gong
authors.forename: Pauline
authors.initials: P
authors.lastname: Fung
authors.forename: Pauline W
authors.initials: PW
authors.lastname: Wang
authors.forename: David S
authors.initials: DS
authors.lastname: Guttman
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics; D016680:Genome, Bacterial; D016366:Open Reading Frames; D044224:Pseudomonas syringae / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial
references.citation: PLoS One. 2011 Jan 27;6(1):e16451
references.pmid: 21304594
references.citation: Bioinformatics. 2011 Feb 15;27(4):578-9
references.pmid: 21149342
references.citation: Genome Res. 2009 Feb;19(2):294-305
references.pmid: 19015323
references.citation: PLoS One. 2010 Jun 25;5(6):e11147
references.pmid: 20593022
references.citation: Trends Genet. 2000 Jun;16(6):276-7
references.pmid: 10827456
references.citation: FEMS Microbiol Lett. 2009 Feb;291(1):103-11
references.pmid: 19077061
references.citation: Mol Plant Microbe Interact. 2009 Jan;22(1):52-62
references.pmid: 19061402
references.citation: BMC Genomics. 2008 Feb 08;9:75
references.pmid: 18261238
references.citation: Environ Microbiol. 2010 Jun;12(6):1604-20
references.pmid: 20370821
references.citation: BMC Genomics. 2009 Aug 24;10:395
references.pmid: 19703286
references.citation: Science. 2009 Oct 9;326(5950):236-7
references.pmid: 19815760
references.citation: PLoS One. 2009 Jul 20;4(7):e6291
references.pmid: 19617911
references.citation: Annu Rev Phytopathol. 2011;49:269-89
references.pmid: 21568703
references.citation: J Bacteriol. 2005 Sep;187(18):6488-98
references.pmid: 16159782
references.citation: PLoS One. 2010 Apr 19;5(4):e10224
references.pmid: 20419105
references.citation: Nucleic Acids Res. 2006 Jan 1;34(Database issue):D344-8
references.pmid: 16381883