Difference between revisions of "Part:BBa K2929004:Design"
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===Design Notes=== | ===Design Notes=== | ||
| − | Besides codon-optimising the sequence for our E. coli chassis, we used a protein tertiary structural predictor to assess if the protein folds correctly. The predictor suggested that the protein folds correctly, having a similar topology to mOrange (PDB ID: 2H5O). This suggested to us that the SpyTag would have no negative impact on the fluorescence of the part. | + | Besides codon-optimising the sequence for our E. coli chassis, we used a protein tertiary structural predictor to assess if the protein folds correctly. The predictor suggested that the protein folds correctly, having a similar topology to mOrange (PDB ID: 2H5O). This suggested to us that the SpyTag would have no negative impact on the fluorescence of the part. |
| + | Added flanking 5' NcoI and 3' BamHI cut sites to allow cloning into our pEHisTEV vector. Added extra Glycine residue 5' to 2xGGSG linker. | ||
===Source=== | ===Source=== | ||
Latest revision as of 15:52, 11 October 2019
mOrange + SpyTag
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 820
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 594
Illegal AgeI site found at 706 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
Besides codon-optimising the sequence for our E. coli chassis, we used a protein tertiary structural predictor to assess if the protein folds correctly. The predictor suggested that the protein folds correctly, having a similar topology to mOrange (PDB ID: 2H5O). This suggested to us that the SpyTag would have no negative impact on the fluorescence of the part.
Added flanking 5' NcoI and 3' BamHI cut sites to allow cloning into our pEHisTEV vector. Added extra Glycine residue 5' to 2xGGSG linker.
Source
Obtained the sequences for BBa_E2050 and BBa_K1159201 from the registry, then joined the sequences via a 2xGGSG linker and ordered sequence from IDT. The sequence was codon-optimised for E. coli using the IDT codon optimisation tool.