ArticleApplied and environmental microbiology2004
Construction and characterization of a highly efficient Francisella shuttle plasmid.
Article in Applied and environmental microbiology, 2004. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 109 papers.
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Who cites it
109 citing papers in PubMed, 155 citations in OpenAlex.
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- Long-Term Survival of Virulent Tularemia Pathogens outside a Host in Conditions That Mimic Natural Aquatic Environments.Applied and environmental microbiology · 2021Article
- Structural and functional analysis of the Francisella lysine decarboxylase as a key actor in oxidative stress resistance.Scientific reports · 2021Article
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- OpiA, a Type Six Secretion System Substrate, Localizes to the Cell Pole and Plays a Role in Bacterial Growth and Viability inJournal of bacteriology · 2020Article
- Tn-Seq reveals hidden complexity in the utilization of host-derived glutathione in Francisella tularensis.PLoS pathogens · 2020Article
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- Efficient transposon mutagenesis mediated by an IPTG-controlled conditional suicide plasmid.BMC microbiology · 2018Article
- Single vector platform vaccine protects against lethal respiratory challenge with Tier 1 select agents of anthrax, plague, and tularemia.Scientific reports · 2018Article
- Mobilizable Plasmids for Tunable Gene Expression inFrontiers in cellular and infection microbiology · 2018Article
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49 more citing papers are in PubMed but not listed here.
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Authors and funding
6 authors at 2 institutions in 2 countries.
Funding
Abstract
Francisella tularensis is a facultative intracellular pathogen that infects a wide variety of mammals and causes tularemia in humans. It is recognized as a potential agent of bioterrorism due to its low infectious dose and multiple routes of transmission. To date, genetic manipulation in Francisella spp. has been limited due to the inefficiency of DNA transformation, the relative lack of useful selective markers, and the lack of stably replicating plasmids. Therefore, the goal of this study was to develop an enhanced shuttle plasmid that could be utilized for a variety of genetic procedures in both Francisella and Escherichia coli. A hybrid plasmid, pFNLTP1, was isolated that was transformed by electroporation at frequencies of >1 x 10(7) CFU mug of DNA(-1) in F. tularensis LVS, Francisella novicida U112, and E. coli DH5alpha. Furthermore, this plasmid was stably maintained in F. tularensis LVS after passage in the absence of antibiotic selection in vitro and after 3 days of growth in J774A.1 macrophages. Importantly, F. tularensis LVS derivatives carrying pFNLTP1 were unaltered in their growth characteristics in laboratory medium and macrophages compared to wild-type LVS. We also constructed derivatives of pFNLTP1 containing expanded multiple cloning sites or temperature-sensitive mutations that failed to allow plasmid replication in F. tularensis LVS at the nonpermissive temperature. In addition, the utility of pFNLTP1 as a vehicle for gene expression, as well as complementation, was demonstrated. In summary, we describe construction of a Francisella shuttle plasmid that is transformed at high efficiency, is stably maintained, and does not alter the growth of Francisella in macrophages. This new tool should significantly enhance genetic manipulation and characterization of F. tularensis and other Francisella biotypes.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.