title: A Tn7-based broad-range bacterial cloning and expression system.
abstract: For many bacteria, cloning and expression systems are either scarce or nonexistent. We constructed several mini-Tn7 vectors and evaluated their potential as broad-range cloning and expression systems. In bacteria with a single chromosome, including Pseudomonas aeruginosa, Pseudomonas putida and Yersinia pestis, and in the presence of a helper plasmid encoding the site-specific transposition pathway, site- and orientation-specific Tn7 insertions occurred at a single attTn7 site downstream of the glmS gene. Burkholderia thailandensis contains two chromosomes, each containing a glmS gene and an attTn7 site. The Tn7 system allows engineering of diverse genetic traits into bacteria, as demonstrated by complementing a biofilm-growth defect of P. aeruginosa, establishing expression systems in P. aeruginosa and P. putida, and 'GFP-tagging' Y. pestis. This system will thus have widespread biomedical and environmental applications, especially in environments where plasmids and antibiotic selection are not feasible, namely in plant and animal models or biofilms.
authors.affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado 80523, USA.
authors.forename: Kyoung-Hee
authors.initials: KH
authors.lastname: Choi
authors.forename: Jared B
authors.initials: JB
authors.lastname: Gaynor
authors.forename: Kimberly G
authors.initials: KG
authors.lastname: White
authors.forename: Carolina
authors.initials: C
authors.lastname: Lopez
authors.forename: Catharine M
authors.initials: CM
authors.lastname: Bosio
authors.forename: RoxAnn R
authors.initials: RR
authors.lastname: Karkhoff-Schweizer
authors.forename: Herbert P
authors.initials: HP
authors.lastname: Schweizer
mesh_terms: D002876:Chromosomes, Bacterial / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D004251:DNA Transposable Elements / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis*
chemical_list: D004251:DNA Transposable Elements; D011994:Recombinant Proteins