Development of a synthetic cumate-inducible gene expression system for Bacillus

Seo, Seung Beom; Schmidt‐Dannert, Claudia

doi:10.1007/s00253-018-9485-4

Cited by 35 publications

(41 citation statements)

References 33 publications

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“…Growth conditions for B. subtilis 168 (referred to as WT) were therefore evaluated using two different media; a Luria Bertani (LB) and a clear, buffered Spizizen minimal medium (SMM). Bacillus cells were transformed with a plasmid (pCT-empty) derived from our previously developed cumate ( p -isopropyl benzoate) inducible Bacillus expression vector ( Table S1 ) 39 . Bacterial growth, media pH, flagella presence, and sporulation were then analyzed for 4 days in 24 h intervals at 20 and 30°C ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Plasmids were constructed using a combination of Gibson Assembly (HiFi® DNA assembly kit, NEB) for fragment assembly, site directed mutagenesis (Q5® mutagenesis kit, NEB) for shorter insertions, deletions and insertions, or restriction enzyme cloning and ligation for larger fragment assembly following manufacturers’ instructions and as described previously 1, 11 .…”

Section: Methodsmentioning

confidence: 99%

“…Briefly, for the C-terminal fusion of biomineralization peptides to His-EutM in pCT5-His-EutM 1, 11 , primers were designed to insert the corresponding into the plasmid using the Q5® mutagenesis kit according to the manufacturers’ instructions.…”

Section: Methodsmentioning

confidence: 99%

“…To create the EutM Bacillus expression vectors, His-EutM-SpyCatcher was amplified from pCT5- His-EutM-SpyC and inserted into our in-house, cumate inducible Bacillus expression vector pCT5-bac2.0 11 by HiFi Assembly. For a control plasmid, pCT-empty was created by deletion of the sfgfp gene from pCT5-bac2.0 using the the Q5® mutagenesis kit according to the manufacturers’ instructions.…”

Section: Methodsmentioning

confidence: 99%

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Engineering Bacillus subtilis for the formation of a durable living biocomposite material

Kang

Pokhrel

Bratsch

et al. 2021

Preprint

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Engineered living materials (ELMs) are a fast-growing area of research that combine approaches in synthetic biology and material science. Here, we engineer B. subtilis to become a living component of a silica material composed of self-assembling protein scaffolds for functionalization and cross-linking of cells. B. subtilis was engineered to display SpyTags on polar flagella for cell attachment and cross-linking of SpyCatcher modified secreted scaffolds. Through deletion of the autolysis LytC, endospore limited B. subtilis cells become a structural component of the material with spores for long-term storage of genetic programming. Known silica biomineralization peptides were screened and scaffolds designed for silica polymerization to fabricate biocomposite materials with enhanced mechanical properties. We show that the resulting ELM can be regenerated from a piece of silica material and that new functions can be readily incorporated by co-cultivation of engineered B. subtilis strains. We believe that this work will serve as a framework for the future design of resilient ELMs as functional, self-healing materials for use as responsive coatings and plasters.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Engineering Bacillus subtilis for the formation of a durable living biocomposite material

Kang

Pokhrel

Bratsch

et al. 2021

Preprint

Self Cite

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“…Seo and Schmidt-Dannert in 2019 used the strong promoter P veg in combination with regulatory elements of Pseudomonas putida to investigate and control the expression of green fluorescent protein (GFP) in B. subtilis. 88 This new system has proved promising and useful for studies of metabolic engineering, synthetic biology, and production of proteins of industrial interest. 88 As a result of numerous studies, the repertoire of techniques has expanded, and improvement of the efficiency of promoters via chromosomal integration has become an option.…”

Section: Impact Statementmentioning

confidence: 99%

The multifunctionality of expression systems in Bacillus subtilis: Emerging devices for the production of recombinant proteins

Souza

Guimarães

Pereira

et al. 2021

Exp Biol Med (Maywood)

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Bacillus subtilis is a successful host for producing recombinant proteins. Its GRAS (generally recognized as safe) status and its remarkable innate ability to absorb and incorporate exogenous DNA into its genome make this organism an ideal platform for the heterologous expression of bioactive substances. The factors that corroborate its value can be attributed to the scientific knowledge obtained from decades of study regarding its biology that has fostered the development of several genetic engineering strategies, such as the use of different plasmids, engineering of constitutive or double promoters, chemical inducers, systems of self-inducing expression with or without a secretion system that uses a signal peptide, and so on. Tools that enrich the technological arsenal of this expression platform improve the efficiency and reduce the costs of production of proteins of biotechnological importance. Therefore, this review aims to highlight the major advances involving recombinant expression systems developed in B. subtilis, thus sustaining the generation of knowledge and its application in future research. It was verified that this bacterium is a model in constant demand and studies of the expression of recombinant proteins on a large scale are increasing in number. As such, it represents a powerful bacterial host for academic research and industrial purposes.

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Improvement of stress tolerance and riboflavin production of Bacillus subtilis by introduction of heat shock proteins from thermophilic bacillus strains

Wang

et al. 2019

Appl Microbiol Biotechnol

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Development of a synthetic cumate-inducible gene expression system for Bacillus

Cited by 35 publications

References 33 publications

Engineering Bacillus subtilis for the formation of a durable living biocomposite material

Engineering Bacillus subtilis for the formation of a durable living biocomposite material

The multifunctionality of expression systems in Bacillus subtilis: Emerging devices for the production of recombinant proteins

Improvement of stress tolerance and riboflavin production of Bacillus subtilis by introduction of heat shock proteins from thermophilic bacillus strains

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