Optimization of GFP levels for analyzing Salmonella gene expression during an infection.
Green fluorescent protein (GFP) is an attractive reporter for Salmonella gene expression analysis but might interfere with virulence when expressed at high levels. To identify suitable GFP levels, we constructed a series of Salmonella strains expressing different amounts of GFP and measured their fluorescence and colonization levels in infected mice. The results show that GFP concentrations in the range of 7000-200,000 molecules per Salmonella cell are detectable in ex vivo samples using flow cytometry, and cause no major Salmonella virulence defect. Appropriate GFP levels can be obtained with weak promoters and stable GFP, or strong promoters and destabilized GFP.
Wendland
Meike
M
Max-Planck-Institute for Infection Biology, Department of Molecular Biology, Schumannstr. 21/22, D-10117 Berlin, Germany.
Bumann
Dirk
D
D000818:Animals; D004195:Disease Models, Animal; D005260:Female; D005434:Flow Cytometry; D015870:Gene Expression; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D012480:Salmonella Infections / Q000382:microbiology; D019779:Salmonella enterica / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity
D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins