Display Settings:

Format

Send to:

Choose Destination
See comment in PubMed Commons below
Genome Res. 2009 Dec;19(12):2308-16. doi: 10.1101/gr.097097.109. Epub 2009 Oct 13.

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

Author information

  • 1The Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Cambridge, United Kingdom.

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.

PMID:
19826075
[PubMed - indexed for MEDLINE]
PMCID:
PMC2792183
Free PMC Article

Images from this publication.See all images (4)Free text

Figure 1.
Figure 2.
Figure 3.
Figure 4.
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Icon for HighWire Icon for PubMed Central Icon for Faculty of 1000
    Loading ...
    Write to the Help Desk