Part:BBa_K3885212
P28-tetR-ssrA
The ssrA degradation tag can degrade the tetR repressor, eliminating repression of downstream genes.
Usage and Biology
The ssrA tag can degrade the abnormal protein during the translation process, so that the tetR gene can be cleaved and lose its expression function, and the original expression of the repressor protein can be degraded. Therefore, we introduced the ssrA tag and constructed part P28-tetR-ssrA.
Characterization
First, we put the part into Cell-Free system to verify its effect. From the fluorescence data, it is clear that our constructed part can function properly. SsrA tag eliminates the effect of residual deterrent proteins in the system. In the follow-up experiment, in order to further improve the degradation effect of ssrA tag, we introduced ClpXP which can accelerate the degradation of the deterrent protein and verified its binding effect.
Design Page
We found that even after the Cas9 protein exercises cleavage activity to disable tetR, repressor proteins remain in the system still prevent fluorescent protein expression and have an impact on the signaling output of the system . Therefore, by reviewing the literature, we identified the ssrA degradation tag. ssrA tags are encoded by amino acids that can be added to the C-terminus of proteins to achieve degradation of abnormal proteins during translation.Thus, by adding the ssrA tag to the back end of tetR, we were able to solve the above problem.
References
Marshall R, Noireaux V. Synthetic Biology with an All E. coli TXTL System: Quantitative Characterization of Regulatory Elements and Gene Circuits. Methods Mol Biol. 2018;1772:61-93.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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