Part:BBa_K2087001:Design

atoxic C. difficile toxin B

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal SpeI site found at 5300
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 1589
  Illegal SpeI site found at 5300
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal SpeI site found at 5300
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal SpeI site found at 5300
  Illegal NgoMIV site found at 7154
  
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

Quite a large protein, so normal E. coli strains might not be able to fold it properly. B. megaterium was used to produce atoxic C. diffile toxin B, so it would be a better bet to produce this protein. Additionally, the size of the linker may be optimized for the desired domain to be delivered.

There is a Spe1 cut site inside the aTcdB gene, so another pair of restriction enzymes should be used when cloning from the biobrick.

Source

An atoxic mutant of the C. difficile toxin B gene.

References

Krautz-Peterson G, Zhang Y, Chen K, Oyler GA, Feng H, Shoemaker CB. Retargeting Clostridium difficile Toxin B to neuronal cells as a potential vehicle for cytosolic delivery of therapeutic biomolecules to treat botulism. J. Toxicol. (2012)