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pCold DNA Cold Shock Expression System

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Takara’s pCold Expression Vectors offer cold shock expression technology for high purity, high yield protein production.

 

The pCold series includes several different vectors. Each includes the cold shock Protein A (cspA) promoter for expression of high purity, high yield recombinant protein in E. coli. These vectors selectively induce target protein synthesis at a low temperature (15°C), a condition which suppresses the expression of host proteins and decreases protease activity. This results in high yields of target proteins (60% of intracellular protein). In addition to the cspA promoter, all vectors contain a lacoperator, ampicillin resistance gene (ampr), ColE1 origin of replication, M13 IG fragment, and a multiple cloning site (MCS). Four of the vectors contain either a translation enhancing element (TEE), His-Tag sequence, and/or Factor Xa cleavage site. The pCold GST vector contains a glutathione S-transferase (GST) sequence and can be used for the expression of proteins with a N-terminal GST tag. 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.


Technical Data:

Features

  • Expression of difficult proteins that can not be expressed with the T7 system
  • Increased solubility due to expression at reduced temperature
  • Increased purity due to repressed expression of host proteins
  • When used in conjunction with one of Takara’s Chaperone Plasmids, the amount of recoverable soluble protein can be further increased
  • Radioisotope Labeling: Up to 90% of newly expressed cellular protein is labeled target protein
  • Chain Length:
    • pCold I DNA: 4,407 bp
    • pCold II DNA: 4,392 bp
    • pCold III DNA: 4,377 bp
    • pCold IV DNA: 4,359 bp
    • pCold GST DNA: 5,097 bp

Applications

  • High-efficiency protein expression using a cold shock promoter

Purity

  • Agarose gel electrophoresis indicates over 70% double-stranded covalently closed circular DNA (RF I).
  • Maintenance of cloning sites was confirmed.
  • Single site cleavage by restriction enzymes Nde I, Sac I, Kpn I, Xho I, BamH I, EcoR I, Hind I, Sal I, Pst I and Xba I was confirmed.

Storage

–20°C

pCold Vector GenBank Accession Numbers

Vector GenBank Accession No.
pCold I DNA AB186388
pCold II DNA AB186389
pCold III DNA AB186390
pCold IV DNA AB186391

Vector Features

Vector TEE His-Tag Factor Xa
Cleavage
Site
pCold I DNA yes yes yes
pCold II DNA yes yes no
pCold III DNA yes no no
pCold IV DNA no no no
pCold GST DNA yes yes yes


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