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a Department of Microbiology of Molecular Genetics, and the Center for Biopreparedness and Infectious Disease, Medical College of Wisconsin, Milwaukee, Wisconsin
Key Words: Francisella • genetics • shuttle vectors • allelic exchange • transposon mutagenesis • complementation
Address for correspondence: Thomas C. Zahrt, Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226–0509. Voice: 414-456-7429; fax: 414-456-6535. tzahrt{at}mcw.edu
Francisella tularensis is a gram-negative coccobacillus and the etiological agent of tularemia. The limited knowledge regarding the interaction of F. tularensis with its host is due in part to the previous lack of tools for genetically manipulating the organism. During the past 10 years, the field of F. tularensis genetics has seen a rapid expansion. Plasmids capable of stable or conditional replication in Francisella have been constructed. Methods for the efficient introduction of DNA into Francisella have been developed or optimized. Genetic platforms and procedures for transposon mutagenesis and allelic exchange have been adapted for use in Francisella. Finally, selectable, counterselectable, and screenable genetic markers amenable for use in a variety of F. tularensis species, including highly virulent clinical isolates, have been described. Collectively, these advances have aided in the construction of defined Francisella mutants and helped investigators begin to define the mechanism(s) employed by these organisms to cause disease in the host. In this article, we describe the history of genetic manipulation in Francisella and summarize the current tools and techniques for conducting genetic studies in this organism.
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