Broad-host-range expression vectors that carry the L-arabinose-inducible Escherichia coli araBAD promoter and the araC regulator

We describe the development and analysis of broad-host-range (BHR) cloning vectors that carry the araC-P-BAD controlled expression cassette from Escherichia coli. These plasmids are designed to facilitate L-arabinose-responsive control of target genes in a variety of Gram-negative bacterial hosts. BHR P-BAD:lacZ fusions were used to analyze the utility of this controlled expression system in the plant pathogen Agrobacterium tumefaciens. In A. tumefaciens,the level of control afforded is significant, although less stringent than that observed in E. coli. The BHR P-BAD vectors offer a useful alternative to currently used controlled expression systems, and can be employed in conjunction with other regulated promoters to simultaneously regulate expression of multiple genes. Addition of a variety of carbon sources, namely C4 acids and the anti-inducer D-fucose, allows modulation of L-arabinose induction. Activation of P-BAD expression in A. tumefaciens requires a plasmid-borne copy of araC, and is not affected by endogenous regulators. (C) 1999 Elsevier Science B.V. All rights reserved.