Construction of an efficient expression system for Agrobacterium tumefaciens based on the coliphage T5 promoter

A versatile expression Vector utilizing a promoter of coliphage T5, P-N25 ( Gentz and Bujard, 1985. J. Bacteriol. 164, 70-77) and a derivative of the I ncW broad-host-range plasmid pJB20 (Beaupre et al., 1997. J. Bacteriol. 179 , 78-89) has been developed. This vector successfully expresses virulence p roteins of Agrobacterium tumefaciens encoded by virG and a mutant allele of virA, virA (Delta 1-284, G665D) in Escherichia coli as well as in A. tumef aciens. The signal transduction proteins VirA (Delta 1-284, G665D) and virG are fully functional when expressed in Agrobacterium, and the P-N25 driven expression overrides the complex transcriptional regulation present with t he native promoters. This expression system will enable a more detailed ana lysis of the activation events in signal transduction in A. trumefaciens, a nd we expect it to be useful in other prokaryotes. (C) 2000 Published by El sevier Science B.V. All rights reserved.