Open AccessConstruction of a reporter plasmid for screening in vivo promoter activity in Francisella tularensis
Author(s) -
Kuoppa Kerstin,
Forsberg Åke,
Norqvist Anders
Publication year - 2001
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2001.tb10928.x
Subject(s) - francisella tularensis , chloramphenicol acetyltransferase , biology , plasmid , shuttle vector , chloramphenicol , reporter gene , microbiology and biotechnology , tularemia , francisella , recombinant dna , virology , cloning (programming) , gene , vector (molecular biology) , gene expression , genetics , antibiotics , virulence , computer science , programming language
Francisella tularensis is a facultative intracellular bacterium that survives and multiplies inside macrophages. Here we constructed a new promoter probe plasmid denoted pKK214 by introduction of a promoter‐less chloramphenicol acetyltransferase ( cat ) gene into the shuttle vector pKK202. A promoter library was created in F. tularensis strain LVS by cloning random chromosomal DNA fragments into pKK214. Approximately 15% of the recombinant bacteria showed chloramphenicol resistance in vitro. The promoter library was also used to infect macrophages in the presence of chloramphenicol and after two cycles of infection the library contained essentially only chloramphenicol resistance clones which shows that pKK214 can be used to monitor F. tularensis genes that are expressed during infection.