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Induction of natural competence in Bacillus cereus ATCC14579.

Mirończuk AM, Kovács ÁT, Kuipers OP - Microb Biotechnol (2008)

Bottom Line: Natural competence is the ability of certain microbes to take up exogenous DNA from the environment and integrate it in their genome.When expressing the B. subtilis ComK protein using an IPTG-inducible system in B. cereus ATCC14579, cells grown in minimal medium displayed natural competence, as either genomic DNA or plasmid DNA was shown to be taken up by the cells and integrated into the genome or stably maintained respectively.This work proves that a sufficient structural system for DNA uptake exists in B. cereus.

View Article: PubMed Central - PubMed

Affiliation: Groningen Biomolecular Sciences and Biotechnology Institute, Department of Genetics, University of Groningen, Kerklaan 30, 9751NN Haren, the Netherlands.

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Bacillus subtilis ComK binds to both B. cereus late‐competence genes promoters PcomGABce and PcomEABce (A), and the promoters of comK1 and comK2 genes (B). Assessment of binding of ComKBsu was performed as described in Experimental procedures. DNA fragments from the indicated promoter regions were prepared by PCR and end‐labelled with 33P. In the panel, the lanes marked with an X correspond to probe alone. The remaining lanes correspond to probe samples incubated with ComKBsu, at concentration ranging from 0.2 µM to 1.6 µM. Each lane, from left to right, corresponds to a doubling in the concentration of ComKBsu.
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f4: Bacillus subtilis ComK binds to both B. cereus late‐competence genes promoters PcomGABce and PcomEABce (A), and the promoters of comK1 and comK2 genes (B). Assessment of binding of ComKBsu was performed as described in Experimental procedures. DNA fragments from the indicated promoter regions were prepared by PCR and end‐labelled with 33P. In the panel, the lanes marked with an X correspond to probe alone. The remaining lanes correspond to probe samples incubated with ComKBsu, at concentration ranging from 0.2 µM to 1.6 µM. Each lane, from left to right, corresponds to a doubling in the concentration of ComKBsu.

Mentions: Previously, it has been shown that in B. subtilis ComK can bind directly to a region upstream of late‐competence genes (Hamoen et al., 1998). Expression activation at the comGABsu promoter is solely dependent on ComK (Susanna et al., 2004). Here, electrophoretic mobility shift assays (EMSAs) were performed to determine whether purified ComKBsu protein can bind to the comGAand comE promoter regions of B. cereus(hereafter referred to as PcomGABce and PcomEABce). The results from the gel shift analyses showed that ComKBsu can bind in vitro to both promoter fragments, leading to reduced mobility of the probes (Fig. 4A). These data confirm that ComKBsu may activate transcription at the PcomGABce promoter region by directly binding to an operator region. Reaction mixtures contained poly(dI‐dC) that is known to eliminate non‐specific DNA binding of ComKBsu (Hamoen et al., 1998).


Induction of natural competence in Bacillus cereus ATCC14579.

Mirończuk AM, Kovács ÁT, Kuipers OP - Microb Biotechnol (2008)

Bacillus subtilis ComK binds to both B. cereus late‐competence genes promoters PcomGABce and PcomEABce (A), and the promoters of comK1 and comK2 genes (B). Assessment of binding of ComKBsu was performed as described in Experimental procedures. DNA fragments from the indicated promoter regions were prepared by PCR and end‐labelled with 33P. In the panel, the lanes marked with an X correspond to probe alone. The remaining lanes correspond to probe samples incubated with ComKBsu, at concentration ranging from 0.2 µM to 1.6 µM. Each lane, from left to right, corresponds to a doubling in the concentration of ComKBsu.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3815884&req=5

f4: Bacillus subtilis ComK binds to both B. cereus late‐competence genes promoters PcomGABce and PcomEABce (A), and the promoters of comK1 and comK2 genes (B). Assessment of binding of ComKBsu was performed as described in Experimental procedures. DNA fragments from the indicated promoter regions were prepared by PCR and end‐labelled with 33P. In the panel, the lanes marked with an X correspond to probe alone. The remaining lanes correspond to probe samples incubated with ComKBsu, at concentration ranging from 0.2 µM to 1.6 µM. Each lane, from left to right, corresponds to a doubling in the concentration of ComKBsu.
Mentions: Previously, it has been shown that in B. subtilis ComK can bind directly to a region upstream of late‐competence genes (Hamoen et al., 1998). Expression activation at the comGABsu promoter is solely dependent on ComK (Susanna et al., 2004). Here, electrophoretic mobility shift assays (EMSAs) were performed to determine whether purified ComKBsu protein can bind to the comGAand comE promoter regions of B. cereus(hereafter referred to as PcomGABce and PcomEABce). The results from the gel shift analyses showed that ComKBsu can bind in vitro to both promoter fragments, leading to reduced mobility of the probes (Fig. 4A). These data confirm that ComKBsu may activate transcription at the PcomGABce promoter region by directly binding to an operator region. Reaction mixtures contained poly(dI‐dC) that is known to eliminate non‐specific DNA binding of ComKBsu (Hamoen et al., 1998).

Bottom Line: Natural competence is the ability of certain microbes to take up exogenous DNA from the environment and integrate it in their genome.When expressing the B. subtilis ComK protein using an IPTG-inducible system in B. cereus ATCC14579, cells grown in minimal medium displayed natural competence, as either genomic DNA or plasmid DNA was shown to be taken up by the cells and integrated into the genome or stably maintained respectively.This work proves that a sufficient structural system for DNA uptake exists in B. cereus.

View Article: PubMed Central - PubMed

Affiliation: Groningen Biomolecular Sciences and Biotechnology Institute, Department of Genetics, University of Groningen, Kerklaan 30, 9751NN Haren, the Netherlands.

Show MeSH
Related in: MedlinePlus