Part:BBa_K3202070
PhiC31 Recombinase Site Kit
This composite part is to test the function of recombinase-toxin-bistable system.
In our study, we are aiming to design a bistable system to achieve zero basal expression in the absence of any inducer while ensuring constant transcriptional capacities and induction rates among promoters. The double bistable system combines small RNA-mediated inhibition of translation with the translational coupling of a repressor to the gene of interest. In our system, we have transcriptional repressors tetR expressed through the inducible promoter but translationally inhibited by the MicC sRNA1 respectively. Since RBS2 and RBS3 is directly inhibited by sRNA while they initiates the transcription of the two repressors, the gene of interest is translationally coupled to the repressor gene while three promoters which produces the inhibitory sRNA are targeted by their corresponding repressors. Therefore, repressors and sRNA are mutually inhibitory. Within one single layer of the bistable system, the transcriptional factor is turned on, pBAD initiates the expression of tetR, which raises the concentration of tetR and thereby exerts greater inhibition on PR1 and therefore decreases the concentration of sRNA1. As sRNA1 is inhibited the inhibitory effect it exerts on tetR accordingly decreases, therefore the recombinase is expressed through the first layer, and vice versa. Nevertheless, given that the expression of LacI inhibits PR1, even if the recombinase is expressed PR1 can be flipped over but cannot express the second layer. This composite part is to test the function of the second bistable system and its compatibilty (with toxin encoded within) with the recombinase system.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 276
Illegal BsaI.rc site found at 266
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