Targeted inactivation of Francisella tularensis genes by group II introns

Stephen A. Rodriguez, Jieh Juen Yu, Greg Davis, Bernard P. Arulanandam, Karl E. Klose

Research output: Contribution to journalArticlepeer-review

49 Scopus citations

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 showed that the system works at high efficiency in three different subspecies: F. tularensis subsp. tularensis, F. tularensis subsp. holarctica, and "F. tularensis subsp. 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 contributions of specific genes, and especially those within the FPI, to the pathogenesis of this poorly studied organism.

Original languageEnglish (US)
Pages (from-to)2619-2626
Number of pages8
JournalApplied and Environmental Microbiology
Volume74
Issue number9
DOIs
StatePublished - May 2008

ASJC Scopus subject areas

  • Biotechnology
  • Food Science
  • Applied Microbiology and Biotechnology
  • Ecology

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