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Journal of Bacteriology, December 2002, p. 6572-6580, Vol. 184, No. 23
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.23.6572-6580.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

A Third Transposable Element, ISPpu12, from the Toluene-Xylene Catabolic Plasmid pWW0 of Pseudomonas putida mt-2

Peter A. Williams,* Rheinallt M. Jones, and Linda E. Shaw

School of Biological Sciences, University of Wales Bangor, Bangor, Gwynedd LL57 2UW, Wales, United Kingdom

Received 29 April 2002/ Accepted 9 August 2002

A 3,372-bp insertion sequence, ISPpu12, has been identified on the archetypal toluene-xylene TOL catabolic plasmid pWW0 from Pseudomonas putida mt-2. The insertion sequence element is located on the plasmid between bases 84397 and 87768 in a region which also contains the termini and transposase genes of the catabolic transposons Tn4651 and Tn4653 (A. Greated, L. Lambertson, P. A. Williams, and C. M. Thomas, Environ. Microbiol., in press). ISPpu12 has terminal inverted repeats of 24 bp with three mismatches and contains four open reading frames, a tnpA homologue and three open reading frames (lspA, orf1, and orf2) of undetermined function. After insertion in vitro of a Kmr cassette into ISPpu12 either in the intergenic region between orf1 and orf2 or directly into the orf1 gene and ligation into a suicide vector, the modified ISPpu12-Km transposes at high frequency, often in multiple copies, into the chromosome of a P. putida recipient. Inactivation of lspA, orf1, and orf2 by introducing a 7-bp deletion into the 5' region of each gene had no major effect upon transposition, but a similar mutation of tnpA completely eliminated transposition. Analysis of the literature and of strains derived from the chlorobenzoate-degrading Pseudomonas sp. strain B13 suggests that the promiscuity of this element has played an important role in the history of plasmid pWW0. Database comparisons and the accompanying paper (A. J. Weightman, A. W. Topping, K. E. Hill, L. L. Lee, K. Sakai, J. H. Slater, and A. W. Thomas, J. Bacteriol. 184:6581-6591, 2002) show that ISPpu12 is a transposable element also found in other bacteria.

* Corresponding author. Mailing address: School of Biological Sciences, Memorial Building, University of Wales Bangor, Bangor, Gwynedd LL57 2UW, United Kingdom. Phone: (44) 1248 382363. Fax: (44) 1248 370731. E-mail: p.a.williams{at}bangor.ac.uk.

Journal of Bacteriology, December 2002, p. 6572-6580, Vol. 184, No. 23
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.23.6572-6580.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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