Protein expression vectors for commercial use

Takara's pCold ProS2 DNA is a cold shock expression vector that includes Protein S derived from a Gram-negative myxobacteria Myxococcus xanthus as a solubilization tag. Protein S, (173 aa) is very stable soluble protein present in the spore surface coat of Myxococcus xanthus. Fusing a target gene encoding a protein of interest with the ProS2 Tag, found in pCold ProS2 DNA may induce the stability and solubility of the fusion protein.

The pCold ProS2 DNA Vector consists of the cspA promoter plus additional downstream sequences including a 5'-untranslated region (5' UTR), a translation enhancing element (TEE), a His-Tag sequence, ProS2 Tag, and a multicloning site (MCS). A lac operator is inserted downstream of the cspA promoter to ensure strict regulation of expression. Additionally, recognition sites for HRV 3C Protease, Thrombin, and Factor Xa are located between ProS2 Tag and the Multiple Cloning Site (MCS) and function to facilitate tag removal from the expressed fusion protein. Most E. coli strains can serve as expression hosts. The pCold ProS2 DNA Vector provides cold shock technology for high yield protein expression to facilitate correct protein folding, thus enabling efficient soluble protein production for otherwise intractable target proteins.

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3371: pCold ProS2 DNA

Takara Bio's pCold Expression Vectors offer cold shock expression technology for high purity, high yield protein production. The series includes four different vectors that utilize the cold shock Protein A (cspA) promoter to direct expression of recombinant protein in E. coli. These vectors selectively induce target protein synthesis at low temperatures (15°C) when the synthesis of other proteins is suppressed and protease activity is decreased. This results in high yield of the target protein (60% of intracellular protein). In addition to the cspA promoter, all four vectors (including pCold I) contain a lac operator (for control of expression), ampicillin resistance gene (amp^r), ColE1 origin of replication, M13 IG fragment, and multiple cloning site (MCS). pCold I also contains a Factor Xa cleavage site. These vectors work equally well for synthesis of non-labeled and radiolabeled proteins and can be used in conjunction with Takara Bio's Chaperone Plasmid Set (Cat.# 3340P).

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3361: pCold I DNA

Takara Bio's pCold Expression Vectors offer cold shock expression technology for high purity, high yield protein production. The series includes four different vectors that utilize the cold shock Protein A (cspA) promoter to direct expression of recombinant protein in E. coli. These vectors selectively induce target protein synthesis at low temperatures (15°C) when the synthesis of other proteins is suppressed and protease activity is decreased. This results in high yield of the target protein (60% of intracellular protein). In addition to the cspA promoter, all four vectors contain a lac operator (for control of expression), ampicillin resistance gene (amp^r), ColE1 origin of replication, M13 IG fragment, and multiple cloning site (MCS). These vectors work equally well for synthesis of non-labeled and radiolabeled proteins and can be used in conjunction with Takara Bio's Chaperone Plasmid Set (Cat.# 3340P).

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3362: pCold II DNA

Takara's pCold Expression Vectors offer cold shock expression technology for high purity, high yield protein production. The series includes four different vectors that utilize the cold shock Protein A (cspA) promoter to direct expression of recombinant protein in E. coli. These vectors selectively induce target protein synthesis at low temperatures (15°C) when the synthesis of other proteins is suppressed and protease activity is decreased. This results in high yield of the target protein (60% of intracellular protein). In addition to the cspA promoter, all four vectors contain a lac operator (for control of expression), ampicillin resistance gene (amp^r), ColE1 origin of replication, M13 IG fragment, and multiple cloning site (MCS). Furthermore, pCold III DNA also contains a his tag sequence and a Factor Xa cleavage site. These vectors work equally well for synthesis of non-labeled and radiolabeled proteins and can be used in conjunction with Takara's Chaperone Plasmid Set (Cat.# 3340P).

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3363: pCold III DNA

Takara's pCold Expression Vectors offer cold shock expression technology for high purity, high yield protein production. The series includes four different vectors that utilize the cold shock Protein A (cspA) promoter to direct expression of recombinant protein in E. coli. These vectors selectively induce target protein synthesis at low temperatures (15°C) when the synthesis of other proteins is suppressed and protease activity is decreased. This results in high yield of the target protein (60% of intracellular protein). In addition to the cspA promoter, all four vectors contain a lac operator (for control of expression), ampicillin resistance gene (amp^r), ColE1 origin of replication, M13 IG fragment, and multiple cloning site (MCS). Furthermore, pCold IV DNA contains a translation enhancing element (TEE), his tag sequence, and a Factor Xa cleavage site. These vectors work equally well for synthesis of non-labeled and radiolabeled proteins and can be used in conjunction with Takara's Chaperone Plasmid Set (Cat.# 3340P).

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3364: pCold IV DNA

The pCold TF DNA Vector provides cold shock technology for high yield protein expression combined with Trigger Factor (chaperone) expression to facilitate correct protein folding, thus enabling efficient soluble protein production for otherwise intractable target proteins. pCold TF DNA Vector is a fusion cold shock expression vector that expresses Trigger Factor (TF) chaperone as a soluble tag. Trigger Factor is a prokaryotic ribosome-associated chaperone protein (48 kDa) which facilitates co-translational folding of newly expressed polypeptides. Because of its E. coli origin, TF is highly expressed in E. coli expression systems. The pCold TF DNA Vector consists of the cspA promoter plus additional downstream sequences including a 5' untranslated region (5' UTR), a translation enhancing element (TEE), a His-Tag sequence, and a multicloning site (MCS). A lac operator is inserted downstream of the cspA promoter to ensure strict regulation of expression. Additionally, recognition sites for HRV 3C Protease, Thrombin, and Factor Xa are located between TF-Tag and the Multiple Cloning Site (MCS) and function to facilitate tag removal from the expressed fusion protein. E. coli strains can serve as expression hosts.

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3365: pCold TF DNA

The expression vector pCold GST DNA was developed by incorporating glutathione S-transferase (GST) derived from Schistosoma japonicum as a soluble tag. The proteins expressed using this vector have an N-terminal GST tag, which can improve the stability and solubility of the fused protein. The pCold GST vector includes a 5' untranslated region (5' UTR), translation enhancing element (TEE), his tag, GST tag, and multiple cloning site (MCS) downstream of the cspA promoter. In addition, a lac operator has been inserted downstream of the promoter to allow precise control of gene expression. Finally, because the pCold vector series uses an E. coli promoter, virtually any strain of E. coli can be used as a host for protein expression.

High affinity purification of the GST-tagged fusion proteins expressed using pCold GST is possible. Furthermore, a highly specific HRV 3C protease recognition sequence has been inserted between the GST tag and MCS, allowing removal of the GST tag from the recombinant protein. Because the optimum reaction temperature for HRV 3C protease is low (4–5℃), tag cleavage can be performed under moderate conditions.

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3372: pCold GST DNA

The Chaperone Plasmid Set consists of five different plasmids, each of which is designed to express multiple molecular chaperones that function as a "chaperone team" to enable protein folding. Co-expression of a target protein with one of these chaperone teams increases the recovery of soluble proteins. Each plasmid carries an origin of replication derived from pACYC and a Cm^r gene, which allows use with E. coli expression systems utilizing ColE1-type plasmids containing an ampicillin resistance gene as a marker. The chaperone genes are situated downstream of an araB or Pzt-1 (tet) promoter. Therefore, expression of target proteins and chaperones can be induced individually if the target gene is placed under the control of other promoters (e.g. lac). These plasmids also contain the necessary regulator (araC or tet^r) for each promoter.

Use this set to perform a two step method to construct target/chaperone co-expression systems: In the first step prepare an E. coli host transformed with only the chaperone plasmid. The second step is to prepare competent cells from this new strain and transform this strain with a plasmid expressing the target protein.

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3340: Chaperone Plasmid Set

Bacillus subtilis has become an increasingly popular host for recombinant protein expression. With its ability to secrete protein directly into culture media, amenability to medium- and large-scale fermentation, lack of codon bias, and designation by the U.S. Food and Drug Administration as an organism that is Generally Regarded As Safe (GRAS), it's no wonder that the majority of industrially-produced enzymes are expressed in Bacillus species such as B. subtilis. Optimization of secretion, however, can be necessary to achieve highest yields. To address this, the B. subtilis Secretory Protein Expression System from Takara Bio allows rapid development of a library of B. subtilis clones, each bearing a pBE-S construct in which the ORF for your protein of interest is fused with sequences for 173 unique signal peptides. Perform a downstream assay to identify and select clones which secrete the highest amount of functional protein into the culture media, and you can quickly identify the signal peptide that results in efficient expression of your desired secreted protein.