Part:BBa_K4275012:Design
CipA1B2C
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 314
Illegal XhoI site found at 1237 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 543
Illegal AgeI site found at 325
Illegal AgeI site found at 669
Illegal AgeI site found at 868
Illegal AgeI site found at 1539 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
1. A 6x His affinity purification tag(HHHHHH) added to N-terminal following the start codon, to allow for Ni-NTA purification.
2. The number of cohesins is cut from 9 to 2, and the number of CBM is cut from 2 to 1 to reduce the metabolic burden and increase the probability of successful expression in the E.coli BL21 recombinant protein expression system.
3. DNA sequence is codon-optimized based on the codon-usage table of E.coli Strain K12.MG1655.
Source
Clostridium thermocellum
References
1. Anandharaj, Marimuthu et al. "Constructing A Yeast To Express The Largest Cellulosome Complex On The Cell Surface". Proceedings Of The National Academy Of Sciences, vol 117, no. 5, 2020, pp. 2385-2394. Proceedings Of The National Academy Of Sciences, https://doi.org/10.1073/pnas.1916529117.