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Nat Chem Biol. 2011 Sep 4;7(10):720-9. doi: 10.1038/nchembio.643.

High-frequency transposition for determining antibacterial mode of action.

Author information

1
Infectious Diseases Division, Merck Frosst Center for Therapeutic Research, Kirkland, Quebec, Canada.

Abstract

Connecting bacterial growth inhibitors to molecular targets at the whole-cell level is a major impediment to antibacterial development. Herein we report the design of a highly efficient and versatile bacteriophage-based mariner transposon delivery system in Staphylococcus aureus for determining inhibitor mode of action. Using bacteriophage-mediated delivery of concatameric minitransposon cassettes, we generated nonclonal transposant libraries with genome-wide insertion-site coverage in either laboratory or methicillin-resistant strain backgrounds and screened for drug resistance in situ on a single agar plate in one step. A gradient of gene-target expression levels, along with a correspondingly diverse assortment of drug-resistant phenotypes, was achieved by fitting the transposon cassette with a suite of outward-facing promoters. Using a panel of antibiotics, we demonstrate the ability to unveil not only an inhibitor's molecular target but also its route of cellular entry, efflux susceptibility and other off-target resistance mechanisms.

PMID:
21892185
DOI:
10.1038/nchembio.643
[Indexed for MEDLINE]

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