Applied and Environmental Microbiology 2008-06-01

In vitro mutagenesis of Bacillus subtilis by using a modified Tn7 transposon with an outward-facing inducible promoter.

Christophe Bordi, Bronwyn G Butcher, Qiaojuan Shi, Anna-Barbara Hachmann, Joseph E Peters, John D Helmann

Index: Appl. Environ. Microbiol. 74(11) , 3419-25, (2008)

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Abstract

A Tn7 donor plasmid, pTn7SX, was constructed for use with the model gram-positive bacterium Bacillus subtilis. This new mini-Tn7, mTn7SX, contains a spectinomycin resistance cassette and an outward-facing, xylose-inducible promoter, thereby allowing for the regulated expression of genes downstream of the transposon. We demonstrate that mTn7SX inserts are obtained at a high frequency and occur randomly throughout the B. subtilis genome. The utility of this system was demonstrated by the selection of mutants with increased resistance to the antibiotic fosfomycin or duramycin.