Expression of marker RNAs in Pseudomonas putida

A broad-host-range vector that expresses a unique artificial RNA in Pseudomonas putida has been developed. This vector was derived from the plasmid pBBR1MCS and incorporates regulatory regions from the Escherichia coli ribosomal operon, rrnB. These include the promoters P1 and P2, and the terminators T1 and T2. The gene for the artificial RNA was derived from Vibrio proteolyticus 5S rRNA. The artificial RNA product accumulates to a level that is 10-20% of the total 5S rRNA in P. putida. The RNA product is not incorporated into ribosomes and has a minimal effect on cell growth rate. In contrast, when wild-type V. proteolyticus 5S rRNA was expressed from the vector, it was incorporated into ribosomes. It is expected that this new vector system will allow artificial RNA expression systems to be readily developed for a large variety of species.