pBaSysBioII: an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis

Eric Botella; Mark Fogg; Matthieu Jules; Sjouke Piersma; Geoff Doherty; Annette Hansen; Emma L Denham; Ludovic Le Chat; Patrick Veiga; Kirra Bailey; Peter J Lewis; Jan Maarten van Dijl; Stéphane Aymerich; Anthony J Wilkinson; Kevin M Devine

doi:10.1099/mic.0.035758-0

pBaSysBioII: an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis

Eric Botella, Mark Fogg, Matthieu Jules, Sjouke Piersma, Geoff Doherty, Annette Hansen, Emma L Denham, Ludovic Le Chat, Patrick Veiga, Kirra Bailey, Peter J Lewis, Jan Maarten van Dijl, Stéphane Aymerich, Anthony J Wilkinson, Kevin M Devine

Department of Life Sciences

Research output: Contribution to journal › Article › peer-review

53 Citations (SciVal)

Abstract

Plasmid pBaSysBioII was constructed for high-throughput analysis of gene expression in Bacillus subtilis. It is an integrative plasmid with a ligation-independent cloning (LIC) site, allowing the generation of transcriptional gfpmut3 fusions with desired promoters. Integration is by a Campbell-type event and is non-mutagenic, placing the fusion at the homologous chromosomal locus. Using phoA, murAA, gapB, ptsG and cggR promoters that are responsive to phosphate availability, growth rate and carbon source, we show that detailed profiles of promoter activity can be established, with responses to changing conditions being measurable within 1 min of the stimulus. This makes pBaSysBioII a highly versatile tool for real-time gene expression analysis in growing cells of B. subtilis.

Original language	English
Pages (from-to)	1600-1608
Number of pages	9
Journal	Microbiology
Volume	156
Issue number	6
Early online date	1 Jun 2010
DOIs	https://doi.org/10.1099/mic.0.035758-0
Publication status	E-pub ahead of print - 1 Jun 2010

Keywords

Bacillus subtilis/genetics
Base Sequence
Carbon/metabolism
Gene Expression
Gene Expression Regulation, Bacterial
Green Fluorescent Proteins/genetics
Molecular Sequence Data
Phosphates/metabolism
Plasmids
Promoter Regions, Genetic
Transcription, Genetic

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10.1099/mic.0.035758-0

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Botella, E., Fogg, M., Jules, M., Piersma, S., Doherty, G., Hansen, A., Denham, E. L., Le Chat, L., Veiga, P., Bailey, K., Lewis, P. J., van Dijl, J. M., Aymerich, S., Wilkinson, A. J., & Devine, K. M. (2010). pBaSysBioII: an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis. Microbiology, 156(6), 1600-1608. Advance online publication. https://doi.org/10.1099/mic.0.035758-0

Botella, E, Fogg, M, Jules, M, Piersma, S, Doherty, G, Hansen, A, Denham, EL, Le Chat, L, Veiga, P, Bailey, K, Lewis, PJ, van Dijl, JM, Aymerich, S, Wilkinson, AJ & Devine, KM 2010, 'pBaSysBioII: an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis', Microbiology, vol. 156, no. 6, pp. 1600-1608. https://doi.org/10.1099/mic.0.035758-0

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title = "pBaSysBioII: an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis",

abstract = "Plasmid pBaSysBioII was constructed for high-throughput analysis of gene expression in Bacillus subtilis. It is an integrative plasmid with a ligation-independent cloning (LIC) site, allowing the generation of transcriptional gfpmut3 fusions with desired promoters. Integration is by a Campbell-type event and is non-mutagenic, placing the fusion at the homologous chromosomal locus. Using phoA, murAA, gapB, ptsG and cggR promoters that are responsive to phosphate availability, growth rate and carbon source, we show that detailed profiles of promoter activity can be established, with responses to changing conditions being measurable within 1 min of the stimulus. This makes pBaSysBioII a highly versatile tool for real-time gene expression analysis in growing cells of B. subtilis.",

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doi = "10.1099/mic.0.035758-0",

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AU - Jules, Matthieu

AU - Piersma, Sjouke

AU - Doherty, Geoff

AU - Hansen, Annette

AU - Denham, Emma L

AU - Le Chat, Ludovic

AU - Veiga, Patrick

AU - Bailey, Kirra

AU - Lewis, Peter J

AU - van Dijl, Jan Maarten

AU - Aymerich, Stéphane

AU - Wilkinson, Anthony J

AU - Devine, Kevin M

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AB - Plasmid pBaSysBioII was constructed for high-throughput analysis of gene expression in Bacillus subtilis. It is an integrative plasmid with a ligation-independent cloning (LIC) site, allowing the generation of transcriptional gfpmut3 fusions with desired promoters. Integration is by a Campbell-type event and is non-mutagenic, placing the fusion at the homologous chromosomal locus. Using phoA, murAA, gapB, ptsG and cggR promoters that are responsive to phosphate availability, growth rate and carbon source, we show that detailed profiles of promoter activity can be established, with responses to changing conditions being measurable within 1 min of the stimulus. This makes pBaSysBioII a highly versatile tool for real-time gene expression analysis in growing cells of B. subtilis.

KW - Bacillus subtilis/genetics

KW - Base Sequence

KW - Carbon/metabolism

KW - Gene Expression

KW - Gene Expression Regulation, Bacterial

KW - Green Fluorescent Proteins/genetics

KW - Molecular Sequence Data

KW - Phosphates/metabolism

KW - Plasmids

KW - Promoter Regions, Genetic

KW - Transcription, Genetic

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DO - 10.1099/mic.0.035758-0

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SN - 1350-0872

VL - 156

SP - 1600

EP - 1608

JO - Microbiology

JF - Microbiology

IS - 6

ER -

pBaSysBioII: an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis

Abstract

Keywords

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