Part:BBa_K3606006

PtetO2 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 of the stronger PL promoters in lambda phages, to create a improved promoter with proper expression level while regulated by tetracycline.

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 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:

Under the induction of aTc, the expression level of ptetO2 is very low. We have measured the expression level of PtetO2, PtetO3(BBa_K3606006) 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.

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:

Huanna Qiu, et al. Construction of promoters with tight regulation on chromosome of Escherichia coli. Microbiology China,2018,45(08):1693-1704. (Chinese)