Part:BBa_K4191002:Design
J23119 + B0034 + FAcD + B0010
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 596
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 786
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 73
Illegal BsaI.rc site found at 994
Design Notes
While designing the sequence in SnapGene, we had to remove certain restriction sites (BsaI, Sapl) from our gene sequence (FacD), as the enzymes would digest and reassemble the sequence. Our enzyme sequence was also codon optimized for E. coli, due to its similarity to our model organism P. putida. 20 bases of homology were also added to the end of the gene sequence prior to ordering the sequence from IDT, to ensure maximum efficiency. In order to ensure the sequence was compatible with our u loop system, the overhangs were modified to match with the C-D overhangs specifically for loop assembly. The other iGEM designed parts had their overhangs modified to fit the respective A-B (promoter), B-C (RBS), and D-F (terminator) overhangs for the u loop system.
Source
This part is composed of a promoter (BBa_J23119), RBS (BBa_B0034), gene sequence (BBa_K4191001), and terminator (BBa_B0010). The gene sequence of fluoroacetate dehalogenase (FAcD) is derived from Rhodopseudomonas palustris and has been codon optimized for E. coli.