This Article Full Text (PDF) Other Versions of this Article: AEM.02905-07v1 74/9/2619    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Rodriguez, S. A. Articles by Klose, K. E. Search for Related Content PubMed PubMed Citation Articles by Rodriguez, S. A. Articles by Klose, K. E. Agricola Articles by Rodriguez, S. A. Articles by Klose, K. E.

 Previous Article  |  Next Article 

Appl. Environ. Microbiol. doi:10.1128/AEM.02905-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Targeted inactivation of Francisella tularensis genes by Group II Introns

South Texas Center for Emerging Infectious Diseases and Department of Biology, University of Texas San Antonio, San Antonio TX 78249; Sigma-Aldrich Biotechnology, Genomics R&D, St. Louis, MO

^* To whom correspondence should be addressed. Email: kklose{at}utsa.edu.

	Abstract

Studies of the molecular mechanisms of pathogenesis of Francisella tularensis, the causative agent of tularemia, have been hampered by a lack of genetic techniques for rapid targeted gene disruption in the most virulent subspecies. Here we describe efficient targeted gene disruption in F. tularensis utilizing mobile group II introns (targetrons) specifically optimized for F. tularensis. Utilizing a targetron targeted to blaB, which encodes ampicillin resistance, we show that the system works at high efficiency in three different subspecies: tularensis, holarctica, and novicida. A targetron was also utilized to inactivate F. tularensis subsp. holarctica iglC, a gene required for virulence. The iglC gene is located within the Francisella Pathogenicity Island (FPI), which has been duplicated in the most virulent subspecies. Importantly, the iglC targetron targeted both copies simultaneously, resulting in a strain mutated in both iglC genes in a single step. This system will help illuminate the contribution of specific genes, and especially those within the FPI, to the pathogenesis of this poorly studied organism.