ArticleApplied and environmental microbiology2011
Construction and characterization of a lactose-inducible promoter system for controlled gene expression in Clostridium perfringens.
Article in Applied and environmental microbiology, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 44 papers.
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44 citing papers in PubMed, 93 citations in OpenAlex.
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- Changes in the expression of genes encoding type IV pili-associated proteins are seen when Clostridium perfringens is grown in liquid or on surfaces.BMC genomics · 2020Article
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3 authors at 1 institution in 1 country.
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Abstract
Clostridium perfringens is a Gram-positive anaerobic pathogen which causes many diseases in humans and animals. While some genetic tools exist for working with C. perfringens, a tightly regulated, inducible promoter system is currently lacking. Therefore, we constructed a plasmid-based promoter system that provided regulated expression when lactose was added. This plasmid (pKRAH1) is an Escherichia coli-C. perfringens shuttle vector containing the gene encoding a transcriptional regulator, BgaR, and a divergent promoter upstream of gene bgaL (bgaR-P(bgaL)). To measure transcription at the bgaL promoter in pKRAH1, the E. coli reporter gene gusA, encoding β-glucuronidase, was placed downstream of the P(bgaL) promoter to make plasmid pAH2. When transformed into three strains of C. perfringens, pAH2 exhibited lactose-inducible expression. C. perfringens strain 13, a commonly studied strain, has endogenous β-glucuronidase activity. We mutated gene bglR, encoding a putative β-glucuronidase, and observed an 89% decrease in endogenous activity with no lactose. This combination of a system for regulated gene expression and a mutant of strain 13 with low β-glucuronidase activity are useful tools for studying gene regulation and protein expression in an important pathogenic bacterium. We used this system to express the yfp-pilB gene, comprised of a yellow fluorescent protein (YFP)-encoding gene fused to an assembly ATPase gene involved in type IV pilus-dependent gliding motility in C. perfringens. Expression in the wild-type strain showed that YFP-PilB localized mostly to the poles of cells, but in a pilC mutant it localized throughout the cell, demonstrating that the membrane protein PilC is required for polar localization of PilB.
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