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Development of a synthetic cumate-inducible gene expression system for Bacillus.
- Source :
-
Applied microbiology and biotechnology [Appl Microbiol Biotechnol] 2019 Jan; Vol. 103 (1), pp. 303-313. Date of Electronic Publication: 2018 Nov 03. - Publication Year :
- 2019
-
Abstract
- A novel inducible gene expression system using p-isopropyl benzoate (cumate) as an inducer was developed for the industrial production hosts, Bacillus subtilis and Bacillus megaterium. Cumate is non-toxic to the host, inexpensive, and carbon source-independent inducer which provides an economical option for large-scale production of valuable proteins and chemicals from Bacillus strains. The synthetic cumate-inducible system was constructed by combining the strong constitutive Bacillus promoter P <subscript>veg</subscript> with regulatory elements of the Pseudomonas putida, CymR repressor, and its operator sequence CuO. The designed expression cassette containing a sfGFP reporter under the cumate-inducible promoter was assembled into a Bacillus-E. coli shuttle and gene expression investigated in the two Bacillus strains. Characterization of gene expression levels, expression kinetics, and dose-response to cumate inducer concentration confirmed high-level, but tightly controlled GFP reporter expression in tunable, cumate concentration-dependent manner. Unexpectedly, this expression system works equally well for Escherichia coli, resulting in a platform that can be used both in gram-positive and gram-negative expression host. Its tight regulation and controllable expression makes this system useful for metabolic engineering, synthetic biology studies as well industrial protein production.
- Subjects :
- Bacillus megaterium drug effects
Bacillus subtilis drug effects
Benzoates administration & dosage
Escherichia coli genetics
Gene Expression Profiling
Genetic Vectors
Green Fluorescent Proteins genetics
Microorganisms, Genetically-Modified
Plasmids genetics
Promoter Regions, Genetic
Pseudomonas putida genetics
Regulatory Sequences, Nucleic Acid
Bacillus megaterium genetics
Bacillus subtilis genetics
Benzoates pharmacology
Gene Expression Regulation, Bacterial drug effects
Genetic Engineering methods
Subjects
Details
- Language :
- English
- ISSN :
- 1432-0614
- Volume :
- 103
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Applied microbiology and biotechnology
- Publication Type :
- Academic Journal
- Accession number :
- 30392122
- Full Text :
- https://doi.org/10.1007/s00253-018-9485-4