cobA, a red fluorescent transcriptional reporter for Escherichia coli, yeast, and mammalian cells.
We demonstrate the use of Propionibacterium freudenreichii uroporphyrinogen III methyltransferase (cobA) as a reporter of gene expression in Escherichia coli, fission yeast, and mammalian cells. Overexpression of cobA in cells resulted in bright red fluorescence that was visualized with standard fluorescence microscopy and fluorescence-activated cell sorting analysis at the single-cell level. As with green fluorescent protein (GFP), no addition of exogenous substrate was required. When expressed in Chinese hamster ovary cells from a bicistronic transcript, cobA and GFP gave rise to fluorescence signals of similar intensity. The bright red fluorescence generated by the cobA reporter promises a better signal-to-noise ratio than blue and green fluorescent reporter systems, as autofluorescence and light scattering of cells, media, and materials are reduced in the red wavelengths.
Wildt
S
S
Pharma Division, Preclinical CNS Research and GeneTechnologies, Molecular Neurobiology, F. Hoffmann-La Roche AG, Basel, Switzerland.
Deuschle
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U
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics*; D005453:Fluorescence; D017930:Genes, Reporter; D008780:Methyltransferases / Q000235:genetics*; D012568:Schizosaccharomyces / Q000235:genetics*; D014158:Transcription, Genetic
D017931:DNA Primers; D008780:Methyltransferases; C060508:uroporphyrin-III C-methyltransferase