1887

Abstract

An unstable type of green fluorescent protein (Gfp) tagged with a C-terminal extension, which is a target for tail-specific protease, was used as a reporter gene in . To analyse Gfp expression in legionellae, transcriptional fusions of unstable with the specific (ntraellular ultiplication) promoters (P, P and P) were constructed. Infection studies using J774.1 macrophages as the host, and strains carrying P-, P- and P- fusions, indicated that the , and genes could be expressed intracellularly. Expression of , and genes in infected cells was examined by flow cytometry. Furthermore, fluorescent intracellular legionellae were detected directly by confocal microscopy. Real-time quantitative RT-PCR revealed the differences in the gene expression of , and that of and , during infection. Expression of was high in the late stage of infection, while that of and was high in the early phase only. We show that unstable is a useful reporter gene whose expression in legionellae can be followed in real-time, and that it allows analysis of promoter activities in legionellae and monitoring of the infection process.

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2008-04-01
2020-08-15
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