Use of a halobacterial reporter gene to analyse the regulation of gene expression in halophilic archaea Free

Abstract

The reading frame encoding a β-galactosidase of ‘’ was used as a reporter gene to investigate three different promoter regions derived from genes of (mc-) and (c- and p-) in transformants. The fusion of at the start codon of each reading frame (A1– fusion genes) caused translational problems in some cases. Transformants containing constructs with fusions further downstream in the reading frame (A–) produced β-galactosidase, and colonies on agar plates turned blue when sprayed with X-Gal. The β-galactosidase activities quantified by standard ONPG assays correlated well with the mRNA data determined with transformants containing the respective genes: the cA– fusion gene was completely inactive, the mcA– transformants showed low amounts of products, whereas the pA– fusion gene was constitutively expressed in the respective transformants. The transcription of each A– gene was activated by the homologous transcriptional activator protein GvpE. The cGvpE, pGvpE and mcGvpE proteins were able to activate the promoter of pA– and mcA–, whereas the promoter of cA– was only activated by cGvpE. Among the three GvpE proteins tested, cGvpE appeared to be the strongest transcriptional activator.

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2001-07-01
2024-03-29
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