Construction of targeted insertion mutations in Francisella tularensis subsp. novicida

Jirong Liu, Xhavit Zogaj, Jeffrey R. Barker, Karl E. Klose

Research output: Contribution to journalArticlepeer-review

27 Scopus citations

Abstract

Francisella tularensis is one of the most deadly bacterial agents, yet most of the genetic determinants of pathogenesis are still unknown. We have developed an efficient targeted mutagenesis strategy in the model organism F. tularensis subsp. novicida by utilizing universal priming of optimized antibiotic resistance cassettes and splicing by overlap extension (SOE). This process enables fast and efficient construction of targeted insertion mutations in F. tularensis subsp. novicida that have characteristics of nonpolar mutations; optimized targeted mutagenesis strategies will promote the study of this mysterious bacterium and facilitate vaccine development against tularemia. Moreover, the general strategy of gene disruption by PCR-based antibiotic resistance cassette insertion is broadly applicable to many bacterial species.

Original languageEnglish (US)
Pages (from-to)487-492
Number of pages6
JournalBioTechniques
Volume43
Issue number4
DOIs
StatePublished - Oct 2007

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)

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