Part:BBa_K2933194

T7 promoter+RBS b+linker h+His+Linker a+Sumo+Linker b+BCII-194+T7 terminator

The part consists of T7 promoter,RBS and protein coding(His+Linker a+Sumo+Linker b+BCII-194)and the biological module can be built into E.coil for protein expression.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 390
  Illegal XbaI site found at 47
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 390
  Illegal NheI site found at 167
  Illegal NheI site found at 1268
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 390
  Illegal BglII site found at 279
  Illegal BamHI site found at 478
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 390
  Illegal XbaI site found at 47
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 390
  Illegal XbaI site found at 47
  
• 1000
  COMPATIBLE WITH RFC[1000]

Usage and Biology

This composite part is made up with nine basic parts, T7 promoter, the RBS b， the linker h， His tag，the linker a, Sumo tag, linker b, the gene of Bcll-194 and T7 terminator.It encodes a protein which is Bcll-194 fused with His and Sumo tag. The fusion protein is about 40.1 kD. In order to gain the highly purified target protein, we add GST tag in N-terminal of Bcll-194 and combine Sumo tag to increased protein solubility. The fusion protein can be cut off at the cutting site by Prescission Protease. It is convenient for us to purify our target protein.

Molecular cloning

First,we obtained BcII by PCR.

Figure 1. The PCR result of BcII.

References

[1]Sung-Kun Kim, Mara Demuth, Sara R. Schlesinger, Sung Joon Kim, Jonathan Urbanczyk, Robert W. Shaw & Hyunshun Shin (2016) Inhibition of Bacillusanthracis metallo-β- lactamase by compounds with hydroxamic acid functionality, Journal of Enzyme Inhibition and Medicinal Chemistry, 31:sup4, 132-137, DOI: 10.1080/14756366.2016.1222580