Composite

Part:BBa_K3202051

Designed by: Heyuan Ni   Group: iGEM19_BHSF_ND   (2019-10-16)
Revision as of 17:41, 16 October 2019 by Registry (Talk | contribs)

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AraC-Pc-pBAD-TetR-RBS2-sfGFP-T500-T1/TE-MicCsRNA-sRNA Binding Site-PLtetO

This composite part is a improved version of the parts BBa_K3202046 and BBa_K3202047 which presents a bistble system; B0034 is replaced with a RBS. Alternatives are BBa_K3202048, BBa_K3202049, and BBa_K3202050 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.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1144
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 979
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1856
    Illegal SapI site found at 961
    Illegal SapI.rc site found at 1879


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Categories
Parameters
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