Part:BBa_K2933144
His+Linker f+AFM-1
This part encodes the fusion protein of His tag and SPG-1 to promote the expression and purification of target protein(AFM-1).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 51
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 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 AFM-1. It encodes a protein which is AFM-1 fused with His tag. The fusion protein is about 28.2 kD. It is convenient for us to purify our target protein.
Molecular cloning
First, we used the vector pET28Ba to construct our expression plasmid. And then we converted the plasmid constructed to E. coli DH5α to expand the plasmid largely.
Figure 1. Left: The PCR result of AFM-1. Right: 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.
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