Gene targeting in gram-negative bacteria by use of a mobile group II intron ("Targetron") expressed from a broad-host-range vector |
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Authors: | Yao Jun Lambowitz Alan M |
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Institution: | Institute for Cellular and Molecular Biology, University of Texas at Austin, 1 University Station A4800, 2500 Speedway, Austin, TX 78712, USA. |
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Abstract: | Mobile group II introns ("targetrons") can be programmed for insertion into virtually any desired DNA target with high frequency and specificity. Here, we show that targetrons expressed via an m-toluic acid-inducible promoter from a broad-host-range vector containing an RK2 minireplicon can be used for efficient gene targeting in a variety of gram-negative bacteria, including Escherichia coli, Pseudomonas aeruginosa, and Agrobacterium tumefaciens. Targetrons expressed from donor plasmids introduced by electroporation or conjugation yielded targeted disruptions at frequencies of 1 to 58% of screened colonies in the E. coli lacZ, P. aeruginosa pqsA and pqsH, and A. tumefaciens aopB and chvI genes. The development of this broad-host-range system for targetron expression should facilitate gene targeting in many bacteria. |
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