Part:BBa_K4451004:Design
TadA*-T7RNAP
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 499
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 2448
Illegal NheI site found at 2496
Illegal PstI site found at 499 - 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 499
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 499
Illegal NgoMIV site found at 1288 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
TadA* has previously been used as a T7RNAP fusion by Evry Paris-Saclay 2021 as part of their Evolution.T7 system (BBa_K3766004). BBa_K4451004 differs slightly in that this part has been codon-optimised for expression in Vibrio natriegens, a marine bacterium that is capable of shorter doubling times than E. coli. Sheffield 2022 showed that due to the faster doubling time of V. natriegens, one would expect fixation of beneficial alleles to occur at a faster rate than E. coli (https://2022.igem.wiki/sheffield/model).
BBa_K4451004 was assembled into pJKR-L under the control of an anhydrotetracycline-inducible promoter (BBa_K4451019), but unfortunately this part could not be tested alongside BBa_K4451022 as a ‘rEvolver’ proof-of-concept before the project deadline.
Source
https://doi.org/10.1038/s41587-020-0491-6
References
Gaudelli, N. M. et al. Directed evolution of adenine base editors with increased activity and therapeutic application. Nat. Biotechnol. 38, 892–900 (2020).
Moore, C. L., Papa, L. J. & Shoulders, M. D. A Processive Protein Chimera Introduces Mutations across Defined DNA Regions In Vivo. J. Am. Chem. Soc. 140, 11560–11564 (2018).