Part:BBa_K524201
Modified ccdB for plasmid assembly on standard vectors
This part was modified from the existing BioBrick BBa_P1010 (ccdB cell death gene), flanked by restriction sites AvrII (followed after RFC10 prefix) and NheI (preceding RFC10 suffix).
One of the many ways to construct plasmids from BioBricks would be to perform standard assembly on a standard vector, excise the insert out using XbaI and SpeI, and then allow the linearized plasmid to self ligate. Such strategy usually involves the purification of desired fragment by electrophoresis followed by gel purification. If the insert is larger than the standard vector, the excised gel fragment might be contaminated by the vector. Contaminant standard vectors would self-ligate and possess higher transformation efficiency than the desired constructed plasmid. In cases where such 'co-transformation' are not desired, the modified ccdB cell death gene can be added at either end of insert by standard assembly (in such situation a ccdB tolerant strain should be used to harbor the construct), and the intended fragment can be excised using either a combination of XbaI + AvrII or SpeI + NheI. As transformation proceeds, any undesired fragments carrying the ccdB part would kill the host cells and the remaining population would only hold the plasmid of interest.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 682
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 670
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 120
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