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

@article{citeulike:5941394, abstract = {10.1101/gr.097097.109 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 (nsposon irected nsertion-site equencing), we have been able to map 370,000 unique transposon insertion sites to the 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 . 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.}, author = {Langridge, Gemma C. and Phan, Minh-Duy and Turner, Daniel J. and Perkins, Timothy T. and Parts, Leopold and Haase, Jana and Charles, Ian and Maskell, Duncan J. and Peters, Sarah E. and Dougan, Gordon and Wain, John and Parkhill, Julian and Turner, A. Keith}, citeulike-article-id = {5941394}, citeulike-linkout-0 = {http://dx.doi.org/10.1101/gr.097097.109}, citeulike-linkout-1 = {http://genome.cshlp.org/content/early/2009/10/13/gr.097097.109.abstract}, citeulike-linkout-2 = {http://genome.cshlp.org/content/early/2009/10/13/gr.097097.109.full.pdf}, citeulike-linkout-3 = {http://view.ncbi.nlm.nih.gov/pubmed/19826075}, citeulike-linkout-4 = {http://www.hubmed.org/display.cgi?uids=19826075}, day = {13}, doi = {10.1101/gr.097097.109}, issn = {1549-5477}, journal = {Genome Research}, keywords = {essential, salmonella\_typhi, transposon\_insertion}, month = {December}, number = {12}, pages = {2308--2316}, posted-at = {2009-11-09 15:55:13}, priority = {2}, title = {Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants}, url = {http://dx.doi.org/10.1101/gr.097097.109}, volume = {19}, year = {2009} }

TY - JOUR ID - citeulike:5941394 L3 - citeulike-article-id:5941394 N2 - 10.1101/gr.097097.109 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 (nsposon irected nsertion-site equencing), we have been able to map 370,000 unique transposon insertion sites to the 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 . 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. IS - 12 JF - Genome Research SN - 1549-5477 EP - 2316 TI - Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants VL - 19 SP - 2308 KW - essential KW - salmonella_typhi KW - transposon_insertion AU - Langridge, Gemma AU - Phan, Minh-Duy AU - Turner, Daniel AU - Perkins, Timothy AU - Parts, Leopold AU - Haase, Jana AU - Charles, Ian AU - Maskell, Duncan AU - Peters, Sarah AU - Dougan, Gordon AU - Wain, John AU - Parkhill, Julian AU - Turner, Keith PY - 2009/12/13/ UR - http://dx.doi.org/10.1101/gr.097097.109 ER -