Part:BBa_K4806000:Design
CYP3A4 gene for Chlamydomonas reinhardtii (Phytobrick)
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 1104
Illegal PstI site found at 1426
Illegal PstI site found at 1486
Illegal PstI site found at 1958
Illegal PstI site found at 2027
Illegal PstI site found at 2131 - 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 1104
Illegal PstI site found at 1426
Illegal PstI site found at 1486
Illegal PstI site found at 1958
Illegal PstI site found at 2027
Illegal PstI site found at 2131 - 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 1104
Illegal PstI site found at 1426
Illegal PstI site found at 1486
Illegal PstI site found at 1958
Illegal PstI site found at 2027
Illegal PstI site found at 2131 - 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 1104
Illegal PstI site found at 1426
Illegal PstI site found at 1486
Illegal PstI site found at 1958
Illegal PstI site found at 2027
Illegal PstI site found at 2131
Illegal NgoMIV site found at 1348 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
This part has been codon optimized for Chlamydomonas reinhardtii and it is compatible with Phytobricks/MoClo parts.
We had to insert one mutation at 493 bp to fit to the MoClo standards of iGEM. In our lab we are not using the restriction enzyme SapI, but instead BbsI. Therefore, this mutation does not occure in the parts we used in our experiments. This mutation is silent, but it might affect the expression level, since the triplet AGT is much rarer in Chlamydomonas, compared to the triplet AGC, which we used. The other mutations were inserted to reduce the amout of cytosine, improving the synthesis of our DNA and to remove recognition sites of BsaI and BbsI.
Additionally, we had to fuse two guanosine at the c-terminal end of our sequence to stay in reading frame.
Further, we had to incorporate introns because Chlamydomonas reinhardtii is an eukaryotic organism.
Source
https://www.uniprot.org/uniprotkb/P08684/entry