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Recombinant protein expression: Brevibacillus expression systems

Brevibacillus choshinensis is a Gram-positive bacterium well-suited for heterologous protein expression. The Brevibacillus expression system generates secreted target proteins efficiently (Takagi et al. 1989) and are ideal for eukaryotic recombinant protein expression, resulting in a high yield of active protein. The Brevibacillus system is also almost completely free of proteases, allowing for the production of intact protein products.

Brevibacillus choshinensis is a Gram-positive bacterium well-suited for heterologous protein expression. The Brevibacillus expression system generates secreted target proteins efficiently (Takagi et al. 1989) and are ideal for eukaryotic recombinant protein expression, resulting in a high yield of active protein. The Brevibacillus system is also almost completely free of proteases, allowing for the production of intact protein products. Our BIC system involves transfecting in a linear plasmid with a PCR insert and relying on the bacteria to recombine into an expression vector, while the Brevibacillus Expression System II involves first cloning the shuttle vector in E. coli and then transfecting in the complete expression vector.

The Brevibacillus system facilitates disulfide-bond formation, which is often required for activity in proteins of eukaryotic origin. In addition, B. choshinensis serves as an excellent host for intracellular protein production, producing soluble intracellular proteins in the cytoplasm without the formation of inclusion bodies. In fact, Brevibacillus expression systems often work better than comparable E. coli-based systems for the expression of certain recombinant protein targets.

Use of his-tag vectors (pNC-HisE, pNC-HisF, pNC-HisT, pNI-His) allows for quick and easy purification of the expressed target proteins. These tags can be removed by protease treatment following purification.

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Cat. # Product Size License Quantity Details
HB200 ◊Brevibacillus Expression System II 1 Kit USD $1475.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊Brevibacillus Expression System II product License Agreement can be found by clicking here.

The Brevibacillus (Bacillus brevis) Expression System is an efficient system for producing high yields of secreted protein. Using this system, it is possible to produce an unprecedented number of heterologous proteins. The Brevibacillus Expression System is particularly effective in the production of secretory proteins.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB132 ◊pNI-His DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNI-His DNA product License Agreement can be found by clicking here.

pNI-His is a shuttle vector between B. choshinensis and E. coli, as well as an intracellular expression vector. Expression plasmids can be constructed by using E. coli as the host, then transfered into B. choshinensis cells. pNI-His contains the P2 promoter, which does not work in E. coli but works as a strong promoter in B. choshinensis cells, enabling efficient protein production. A his tag sequence is located downstream of the secretion signal peptide and an Enterokinase cleavage site is included so that the his tag can be removed from the fusion protein.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB131 ◊pNI DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNI DNA product License Agreement can be found by clicking here.

pNI is a shuttle vector between B. choshinensis and E. coli, as well as an intracellular expression vector. Expression plasmids can be constructed by using E. coli as the host, then transferred into B. choshinensis cells. pNI contains the P2 promoter, which does not work in E. coli but works as a strong promoter in B. choshinensis cells, enabling efficient protein production.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Vector Map of pNI DNA (Cat. # HB131)

Vector Map of pNI DNA (Cat. # HB131)
Vector Map of pNI DNA (Cat. # HB131).
HB123 ◊pNC-HisE DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNC-HisE DNA product License Agreement can be found by clicking here.

pNC-HisE is a shuttle vector between B. choshinensis and E. coli, as well as a secretory expression vector. Expression plasmids can be constructed by using E. coli as the host, then transferred into B. choshinensis cells. pNC-HisE contains the P2 promoter, which does not work in E. coli but works as a strong promoter in B. choshinensis cells, enabling efficient protein production. A his tag sequence is located downstream of the secretion signal peptide and an Enterokinase cleavage site is included so that the his tag can be removed from the fusion protein.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB122 ◊pNC-HisF DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNC-HisF DNA product License Agreement can be found by clicking here.

pNC-HisF is a shuttle vector between B. choshinensis and E. coli, as well as a secretory expression vector. Expression plasmids can be constructed by using E. coli as the host, then transferred into B. choshinensis cells. pNC-HisF contains the P2 promoter, which does not work in E. coli but works as a strong promoter in B. choshinensis cells, enabling efficient protein production. A his tag sequence is located downstream of the secretion signal peptide and a Factor Xa cleavage site is included so that the his tag can be removed from the fusion protein.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB121 ◊pNC-HisT DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNC-HisT DNA product License Agreement can be found by clicking here.

pNC-HisT is a shuttle vector between B. choshinensis and E. coli, as well as a secretory expression vector. Expression plasmids can be constructed by using E. coli as the host, then transferred into B. choshinensis cells. pNC-HisT contains the P2 promoter, which does not work in E. coli but works as a strong promoter in B. choshinensis cells, enabling efficient protein production. A his tag sequence is located downstream of the secretion signal peptide and a thrombin cleavage site is included so that the his tag can be removed from the fusion protein.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB116 ◊Brevibacillus Competent Cells 10 x 100 uL USD $464.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊Brevibacillus Competent Cells product License Agreement can be found by clicking here.

Brevibacillus Competent Cells are host cells used for transformation with Brevibacillus expression vectors such as pNY326 DNA (Cat. #HB111), pNCMO2 DNA (Cat. #HB112), and pNC-HisT DNA (Cat. #HB121). Brevibacillus Competent Cells are prepared for transformation using the New Tris-PEG (NTP) method for chemical transformation of B. choshinensis strain SP3, which can provide  a transformation efficiency that is approximately equal to that obtained with electroporation. The NTP method also enables direct transformation of a DNA ligation solution without DNA purification.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB116: Brevibacillus Competent Cells

HB116: Brevibacillus Competent Cells
HB114 ◊pNY326-BLA DNA 1 ug USD $252.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNY326-BLA DNA product License Agreement can be found by clicking here.

pNY326-BLA DNA is a positive control plasmid for Brevibacillus secretory production. It contains the gene for Bacillus licheniformis α-amylase (~55 kDa) and produces ≥0.1 g secreted protein/L.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB112 ◊pNCMO2 DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNCMO2 DNA product License Agreement can be found by clicking here.

pNCMO2 is a shuttle vector for Brevibacillus and E. coli. It contains the P2 promoter, which does not work in E. coli but works as a strong promoter in B. choshinensis cells, enabling efficient protein production.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Documents Components You May Also Like
HB111 ◊pNY 326 DNA 10 ug USD $443.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pNY 326 DNA product License Agreement can be found by clicking here.

pNY326 can replicate in B. choshinensis cells and its size (3.4 kb) is much smaller than that of pNCMO2 (5.2 kb). In addition, the promoter activity of the vector is substantially weaker than that of pNCMO2. It can be used to express toxic proteins. pNY326 exhibits stable productivity and is especially suited to large scale culture. The expression plasmid must be constructed in one step using B. choshinensis as the host. Because the transformation efficiency must be high, the use of Brevibacillus Competent Cells (105 transformants/μg DNA; Cat. #HB116) is recommended.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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HB300 ◊BIC System Each USD $1476.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊BIC System product License Agreement can be found by clicking here.

The Brevibacillus Expression System is an efficient system for producing secreted proteins and is ideal for eukaryotic recombinant protein expression. Brevibacillus choshinensis is a gram-positive bacterium well-suited for heterologous protein expression and is capable of producing high yields of active protein.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Overview of the Brevibacillus in vivo Cloning (BIC) method

Overview of the Brevibacillus in vivo Cloning (BIC) method
Overview of the Brevibacillus in vivo Cloning (BIC) method.

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pBIC vector map

pBIC vector map
pBIC vector map.
HB310 ◊pBIC DNA Set 1 Kit USD $1053.00

License Statement

ID Number  
L39 1. Brevibacillus Expression System-related products were developed and manufactured by Higeta Shoyu Co., Ltd. and sold by TAKARA BIO INC.
2. The use of this product is allowed for research use only.
3. All rights of intellectual property related to this product are owned by Higeta Shoyu Co., Ltd. A copy of the ◊pBIC DNA Set product License Agreement can be found by clicking here.

The Brevibacillus Expression System is an efficient system for producing secreted proteins and is ideal for eukaryotic recombinant protein expression. Brevibacillus choshinensis is a gram-positive bacterium well-suited for heterologous protein expression and is capable of producing high yields of active protein.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Overview

  • Efficient production of secreted or intracellular recombinant target proteins
  • Negligible amounts of extracellular protease—products remain intact in culture medium
  • Unlike E. coli, no endotoxins
  • Proteins are produced in active form
  • Easy to culture, handle, and sterilize

More Information

Components

Components for the expression systems are provided in their respective user manuals.

Note: This product requires completion of a License Agreement before purchase.

Examples of proteins expressed using the Brevibacillus Expression System II

Examples of expressed proteins using the Brevibacillus Expression System II are shown in the table below. High expression levels have been achieved for a variety of proteins (enzymes, antigens, and cytokines) regardless of their genetic origin (bacterial, archaeal, or eukaryotic). Because secreted eukaryotic proteins often depend on intact disulfide bonds for activity, it is generally difficult to produce active proteins using typical prokaryote-based expression systems. However, due to the secretory advantages of the Brevibacillus Expression System II, high expression levels of active recombinant protein are possible even for proteins with extensive disulfide bonds.

Proteins Origins Production (g/l) Product citations
Enzymes
α-amylase B. licheniformis 3.7
Sphingomyelinase B. cereus 3.0
Xylanase B. halodurans 0.2
CGTase B. macerans 1.5 Yamamoto et al. 2011
Chitosanase B. circulans 1.4
Hyper thermo-stable protease A. pernix 0.1
Hyper thermo-stable nuclease P. horikoshii 0.7
PDI Human 1.0 Sugimoto et al. 2011
Antigens
Surface antigen E. rhusiopathiae 0.9
Surface antigen T. pallidum 0.8
Cytokines
EGF Human 1.5 Mizukami et al. 2010
NGF Mouse 0.2
IFN-γ Chicken 0.5 Yamamoto et al. 2009
TNF-α Bovine 0.4
GM-CSF Bovine 0.2
GH Flounder 0.2

   

References

Takagi, H., Kadowaki, K. & Udaka, S. Screening and characterization of protein-hyperproducing bacteria without detectable exoprotease activity. Agric. Biol. Chem. 53, 691–699 (1989).

Product citations

Mizukami, M., Hanagata, H. & Miyauchi, A. Brevibacillus Expression System: Host-Vector System for Efficient Production of Secretory Proteins. Curr. Pharm. Biotechnol. 11, 251–258 (2010).

Sugimoto, S. et al. Cloning, expression and purification of extracellular serine protease Esp, a biofilm-degrading enzyme, from Staphylococcus epidermidis. J. Appl. Microbiol. 111, 1,406–1,415 (2011).

Takano, T. et al. Expression of the Cycledextrin Glucanotransferase Gene of Bacillus macerans in Bacillus brevis. Biosci. Biotech. Biochem. 56, 808–809 (1992).

Teramura, N. et al. Cloning of a Novel Collagenase Gene from the Gram-Negative Bacterium Grimontia (Vibrio) hollisae 1706B and Its Efficient Expression in Brevibacillus choshinensis. J. Bacteriol. 193, 3,049–3,056 (2011).

Tojo, H. et al. Production of human protein disulfide isomerase by Bacillus brevis. J. Biotechnol. 33, 55-62 (1994).

Yamagata, H., Nakahama, N., Suzuki, Y., Tsukagoshi, N. & Udaka, S. Use of Bacillus brevis for efficient synthesis and secretion of human epidermal growth factor. Proc. Natl. Acad. Sci. USA. 86, 3,589–3,593 (1989).

Yamamoto, A., Fujino, M., Tsuchiya, T. & Iwata, A. Recombinant canine granulocyte colony-stimulating factor accelerates recovery from cyclophosphamide-induced neutropenia in dogs. Vet. Immunol. Immunopathol. 142, (2011).

Yamamoto, A., Iwata, A., Saito, T., Watanabe, F. & Ueda, S. Expression and purification of canine granulocyte colony-stimulating factor (cG-CSF). Vet. Immunol. Immunopathol. 130, 221–225 (2009).

Yashiro, K. et al. High-Level Production of Recombinant Chicken Interferon-γ by Brevibacillus choshinensis. Protein Expr. Purif. 23, 113–120 (2001).

Additional product information

Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.


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Takara Bio USA, Inc. provides kits, reagents, instruments, and services that help researchers explore questions about gene discovery, regulation, and function. As a member of the Takara Bio Group, Takara Bio USA is part of a company that holds a leadership position in the global market and is committed to improving the human condition through biotechnology. Our mission is to develop high-quality innovative tools and services to accelerate discovery.

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