Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants.

Langridge, GC; Phan, M-D; Turner, DJ; Perkins, TT; Parts, L; Haase, J; Charles, I; Maskell, DJ; Peters, SE; Dougan, G; Wain, J; Parkhill, J; Turner, AK

Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants.

Langridge, GC Phan, M-D Turner, DJ Perkins, TT Parts, L Haase, J Charles, I

Maskell, DJ Peters, SE Dougan, G Wain, J Parkhill, J Turner, AK

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Publication Type:: Journal Article
Citation:: Genome Res, 2009, 19 (12), pp. 2308 - 2316
Issue Date:: 2009-12

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Field	Value	Language
dc.contributor.author	Langridge, GC	en_US
dc.contributor.author	Phan, M-D	en_US
dc.contributor.author	Turner, DJ	en_US
dc.contributor.author	Perkins, TT	en_US
dc.contributor.author	Parts, L	en_US
dc.contributor.author	Haase, J	en_US
dc.contributor.author	Charles, I https://orcid.org/0000-0002-2423-1167	en_US
dc.contributor.author	Maskell, DJ	en_US
dc.contributor.author	Peters, SE	en_US
dc.contributor.author	Dougan, G	en_US
dc.contributor.author	Wain, J	en_US
dc.contributor.author	Parkhill, J	en_US
dc.contributor.author	Turner, AK	en_US
dc.date.issued	2009-12	en_US
dc.identifier.citation	Genome Res, 2009, 19 (12), pp. 2308 - 2316	en_US
dc.identifier.uri	http://hdl.handle.net/10453/13343
dc.description.abstract	Very high-throughput sequencing technologies need to be matched by high-throughput functional studies if we are to make full use of the current explosion in genome sequences. We have generated a very large bacterial mutant pool, consisting of an estimated 1.1 million transposon mutants and we have used genomic DNA from this mutant pool, and Illumina nucleotide sequencing to prime from the transposon and sequence into the adjacent target DNA. With this method, which we have called TraDIS (transposon directed insertion-site sequencing), we have been able to map 370,000 unique transposon insertion sites to the Salmonella enterica serovar Typhi chromosome. The unprecedented density and resolution of mapped insertion sites, an average of one every 13 base pairs, has allowed us to assay simultaneously every gene in the genome for essentiality and generate a genome-wide list of candidate essential genes. In addition, the semiquantitative nature of the assay allowed us to identify genes that are advantageous and those that are disadvantageous for growth under standard laboratory conditions. Comparison of the mutant pool following growth in the presence or absence of ox bile enabled every gene to be assayed for its contribution toward bile tolerance, a trait required of any enteric bacterium and for carriage of S. Typhi in the gall bladder. This screen validated our hypothesis that we can simultaneously assay every gene in the genome to identify niche-specific essential genes.	en_US
dc.language	eng	en_US
dc.relation.ispartof	Genome Res	en_US
dc.relation.isbasedon	10.1101/gr.097097.109	en_US
dc.subject.classification	Bioinformatics	en_US
dc.subject.mesh	Chromosomes, Bacterial	en_US
dc.subject.mesh	Bile	en_US
dc.subject.mesh	Salmonella typhi	en_US
dc.subject.mesh	Bacterial Proteins	en_US
dc.subject.mesh	DNA Transposable Elements	en_US
dc.subject.mesh	Chromosome Mapping	en_US
dc.subject.mesh	Mutagenesis, Insertional	en_US
dc.subject.mesh	Sequence Analysis, DNA	en_US
dc.subject.mesh	Computational Biology	en_US
dc.subject.mesh	Mutation	en_US
dc.subject.mesh	Genes, Essential	en_US
dc.title	Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants.	en_US
dc.type	Journal Article
utslib.citation.volume	12	en_US
utslib.citation.volume	19	en_US
utslib.location.activity	United States	en_US
utslib.for	0605 Microbiology	en_US
utslib.for	06 Biological Sciences	en_US
utslib.for	11 Medical and Health Sciences	en_US
dc.location.activity	ISI:000272273400015	en_US
pubs.embargo.period	Not known	en_US
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Strength - ithree - Institute of Infection, Immunity and Innovation
utslib.copyright.status	closed_access
pubs.issue	12	en_US
pubs.publication-status	Published	en_US
pubs.volume	19	en_US

Abstract:

Very high-throughput sequencing technologies need to be matched by high-throughput functional studies if we are to make full use of the current explosion in genome sequences. We have generated a very large bacterial mutant pool, consisting of an estimated 1.1 million transposon mutants and we have used genomic DNA from this mutant pool, and Illumina nucleotide sequencing to prime from the transposon and sequence into the adjacent target DNA. With this method, which we have called TraDIS (transposon directed insertion-site sequencing), we have been able to map 370,000 unique transposon insertion sites to the Salmonella enterica serovar Typhi chromosome. The unprecedented density and resolution of mapped insertion sites, an average of one every 13 base pairs, has allowed us to assay simultaneously every gene in the genome for essentiality and generate a genome-wide list of candidate essential genes. In addition, the semiquantitative nature of the assay allowed us to identify genes that are advantageous and those that are disadvantageous for growth under standard laboratory conditions. Comparison of the mutant pool following growth in the presence or absence of ox bile enabled every gene to be assayed for its contribution toward bile tolerance, a trait required of any enteric bacterium and for carriage of S. Typhi in the gall bladder. This screen validated our hypothesis that we can simultaneously assay every gene in the genome to identify niche-specific essential genes.

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http://hdl.handle.net/10453/13343