title: Sequencing of the Dutch elm disease fungus genome using the Roche/454 GS-FLX Titanium System in a comparison of multiple genomics core facilities.
abstract: As part of the DNA Sequencing Research Group of the Association of Biomolecular Resource Facilities, we have tested the reproducibility of the Roche/454 GS-FLX Titanium System at five core facilities. Experience with the Roche/454 system ranged from <10 to >340 sequencing runs performed. All participating sites were supplied with an aliquot of a common DNA preparation and were requested to conduct sequencing at a common loading condition. The evaluation of sequencing yield and accuracy metrics was assessed at a single site. The study was conducted using a laboratory strain of the Dutch elm disease fungus Ophiostoma novo-ulmi strain H327, an ascomycete, vegetatively haploid fungus with an estimated genome size of 30-50 Mb. We show that the Titanium System is reproducible, with some variation detected in loading conditions, sequencing yield, and homopolymer length accuracy. We demonstrate that reads shorter than the theoretical minimum length are of lower overall quality and not simply truncated reads. The O. novo-ulmi H327 genome assembly is 31.8 Mb and is comprised of eight chromosome-length linear scaffolds, a circular mitochondrial conti of 66.4 kb, and a putative 4.2-kb linear plasmid. We estimate that the nuclear genome encodes 8613 protein coding genes, and the mitochondrion encodes 15 genes and 26 tRNAs.
authors.affiliation: Department of Human Genetics, McGill University, Montréal, Québec, Canada.
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Forgetta
authors.forename: Gary
authors.initials: G
authors.lastname: Leveque
authors.forename: Joana
authors.initials: J
authors.lastname: Dias
authors.forename: Deborah
authors.initials: D
authors.lastname: Grove
authors.forename: Robert
authors.initials: R
authors.lastname: Lyons
authors.forename: Suzanne
authors.initials: S
authors.lastname: Genik
authors.forename: Chris
authors.initials: C
authors.lastname: Wright
authors.forename: Sushmita
authors.initials: S
authors.lastname: Singh
authors.forename: Nichole
authors.initials: N
authors.lastname: Peterson
authors.forename: Michael
authors.initials: M
authors.lastname: Zianni
authors.forename: Jan
authors.initials: J
authors.lastname: Kieleczawa
authors.forename: Robert
authors.initials: R
authors.lastname: Steen
authors.forename: Anoja
authors.initials: A
authors.lastname: Perera
authors.forename: Doug
authors.initials: D
authors.lastname: Bintzler
authors.forename: Scottie
authors.initials: S
authors.lastname: Adams
authors.forename: Will
authors.initials: W
authors.lastname: Hintz
authors.forename: Volker
authors.initials: V
authors.lastname: Jacobi
authors.forename: Louis
authors.initials: L
authors.lastname: Bernier
authors.forename: Roger
authors.initials: R
authors.lastname: Levesque
authors.forename: Ken
authors.initials: K
authors.lastname: Dewar
mesh_terms: D001483:Base Sequence; D016681:Genome, Fungal; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D009181:Mycoses / Q000235:genetics* / Q000382:microbiology; D055156:Ophiostoma / Q000235:genetics*; D010935:Plant Diseases / Q000235:genetics / Q000382:microbiology; D034403:Ulmus / Q000235:genetics* / Q000382:microbiology
keywords: Ophiostoma novo-ulmi; fungal genomics; massively parallel DNA sequencing
references.citation: Can J Microbiol. 2010 Aug;56(8):697-705
references.pmid: 20725133
references.citation: BMC Genomics. 2011 Aug 24;12:431
references.pmid: 21864383
references.citation: Proc Natl Acad Sci U S A. 2011 Feb 8;108(6):2504-9
references.pmid: 21262841
references.citation: J Gen Virol. 1998 Dec;79 ( Pt 12):2921-8
references.pmid: 9880005
references.citation: Proc Natl Acad Sci U S A. 2008 Sep 2;105(35):13081-6
references.pmid: 18723673
references.citation: Nature. 2005 Sep 15;437(7057):376-80
references.pmid: 16056220
references.citation: Genome Res. 2008 Dec;18(12):1979-90
references.pmid: 18757608
references.citation: Genome Res. 2002 Apr;12(4):656-64
references.pmid: 11932250
references.citation: Nucleic Acids Res. 1988 Nov 11;16(21):10393
references.pmid: 3194225
references.citation: Genome Biol. 2009;10(9):R94
references.pmid: 19747388
references.citation: Mol Cell Biol. 1986 Jan;6(1):195-200
references.pmid: 3785146
references.citation: Mol Gen Genet. 1997 Jun;255(1):38-44
references.pmid: 9230897
references.citation: Proc Natl Acad Sci U S A. 1990 Apr;87(7):2833-7
references.pmid: 2320591