Part:BBa_K2933148
His+Linker f+CPS-1
This part encodes the fusion protein of His tag and CPS-1 to promote the expression and purification of target protein(CPS-1).
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 623
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 51
Illegal PstI site found at 623 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 905
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 623
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 623
Illegal AgeI site found at 524 - 1000COMPATIBLE WITH RFC[1000]
Usage and Biology
This composite part is made up with three basic parts, the HiS tag, the thrombin restriction site and our target protein CPS-1. It encodes a protein which is CPS-1 fused with His tag. The fusion protein is about 33.2 kD. It is convenient for us to purify our target protein.
Molecular cloning
First, we used the vector pET28a to construct our expression plasmid. And then we converted the plasmid constructed to E. coli DH5α to expand the plasmid largely.
Figure 1. a: The PCR result of CPS-1. b: The verification results by enzyme digestion.
After verification, it was determined that the construction is successful. We converted the plasmid to E. coli BL21(DE3) for expression and purification.
References
[1]Dereje Dadi Gudeta, Valeria Bortolaia,The Soil Microbiota Harbors a Diversity of Carbapenem-Hydrolyzing β-Lactamases of Potential Clinical Relevance[J]，Antimicrobial Agents and chemotherapy,January 2016
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