Part:BBa_K2669000:Design
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Considerations
To make this part, we had to modify part BBa_K2003011 by moving the start codon to the front of the histidine tag, as opposed to after it. We also wished to express this protein constitutively as opposed to inducibly in order to make it more convenient for us to test if our part worked. In order to prevent potential protein aggregation and/or strain on the cells we expressed our part in a low copy ampicillin backbone ordered from IDT (pUCIDT). In addition, VR and VF2 cloning sites were inserted into our part for convenience, but are not in this composite part.
We also further modified Part:BBa_K2003011 by adding a stop codon (taa) at the end of the flexible GSG linker in order to use it in our composite Part:BBa_K2669000. It is very important to note that this stop codon should be removed/moved to the end of the coding sequence if one wishes to use this part to create a fusion protein expressing construct when using this composite part.
It should be noted that we did not test the flexible linker with another protein, as we did not design the flexible linker ourselves. For more information, please visit the Part:BBa_K2003011 page from Team Uppsala 2016