Regulatory

Part:BBa_K3606007

Designed by: Gaochen Zhang   Group: iGEM20_Fudan   (2020-10-11)
Revision as of 18:36, 25 October 2020 by Cracra (Talk | contribs)


PtetO3 promoter

This part is a fusion promoter of PL and tetO, regulated by tetR and anhydrotetracycline

Background:

As a well-characterized promoter, PtetR has been studied thoroughly with its expression level and leakage.To better optimize the antimicrobial peptide expressing system, here we take a unique idea to created a new promoter fusion to create a new promoter that are regulated by tetracycline while with a desired expression intensity.

Design:

This part is a fusion of tetO operon and the -35 upstream, -35--10 parts，-10 downstream of the stronger PL promoters in lambda phages, to create a improved promoter with proper expression level while regulated by tetracycline. T--Fudan--img_pteto3.png

Usage and Biology:

This part functions to lower the leakage and adjust the level of expression of the downstream gene. We fuse the -35 upstream, -35--10 parts，-10 downstream of the stronger PL promoters in lambda phages with the tetO operon regulated by tetracycline. As a promoter fusion of PL and tetO, not only is it regulated by tetR and anhydrotetracycline, but it also inherited the expression level of PL.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Results:

We have measured the expression level of PtetO2(BBa_K3606007), PtetO3 along with the original PtetR(BBa_R0040) via plate reader by combining them with GFP, when they were induced by different concentration of hydrotetracycline or not induced. Both of these promoters exhibit good low leakage characteristics. Compared to ptetR, their peak intensity can also reach a more proper expression level for our system. 图

Further Application:

This part will act as a tighter tetracycline-regulated promoter with _____ expression level than the original PtetR. The method that applied here can also change the future train of thoughts to create new fusion promoter with different characteristic.

References:

Yingna Qiu, et al. The construction of rigorous type promoter on E. coli chromosomes. Microbiology China,2018,45(08):1693-1704.


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