arabinose-mediated inducible expression of toxin CcdB
This construct is composed of L-arabinose mediated inducible promoter pBAD and toxin CcdB. CcdB/CcdA is a type II toxin-antitoxin system originating from E. coli. In our project, we used this construct to test the toxicity of toxin CcdB, which provided us with more insights in regulating the balance of the toxin-antitoxin system for preventing horizontal gene transfer.
2021 Team Leiden
CcdB has also been used as a part of a kill switch (Fig. 1) to test the toxicity of the toxin based on arabinose concentration.
Figure 1. Plate reader assay of E. coli TOP 10 cellular growth over time with a gradient of arabinose concentrations.
Cell growth is strongly inhibited following the induction of CcdB. The results show that CcdB is not necessarily enough to kill all cells immediately when grown in liquid LB medium. Only after the exponential growth phase, the cells start to die. High concentrations of arabinose are needed for this killing effect, but that might change in different medium conditions.
Sequence and Features
- 10COMPATIBLE WITH RFC
- 12Illegal NheI site found at 1205
- 21Illegal BamHI site found at 1144
Illegal BamHI site found at 1902
- 23COMPATIBLE WITH RFC
- 25Illegal AgeI site found at 979
- 1000Illegal BsaI.rc site found at 1352
Illegal SapI site found at 961