A set of cassettes and improved vectors for genetic and biochemical characterization of Pseudomonas genes

Abstract

A set of broad-host-range vectors allowing direct selection of recombinant DNA molecules to facilitate subcloning and expression analyses of Pseudomonas genes was constructed using Bg/II lacZ alpha cassette. Controlled expression vectors pVDtac39 and pVDtac24 were shown to be useful for determination of enzymatic activities encoded by the cloned DNA fragments and Mr determination of the corresponding polypeptides. A set of Pseudomonas putida xylE gene cassettes truncated at the 5' end was constructed for translational (protein) fusion studies. A protein fusion of the Pseudomonas aeruginosa algD gene, coding for GDPmannose dehydrogenase, and the truncated xylE gene cassette was used to verify the putative coding region and translational signals predicted from the algD nucleotide sequence.