Difference between revisions of "Team:Hong Kong HKUST/pTet"

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<p style="color: black; font-size: 1.2em;"> The TetR repressor exists in 2 states: silent and active. In the absence of inducers, the active state of TetR binds to the TetO1 and TetO2 operator sites of the tetp promoter, thus inhibiting transcription. In the presence of inducers, however, TetR undergoes allosteric rearrangement and switches to its silent state, resulting in its release from the promoter, thus allowing transcription to take place.
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<p style="color: black; font-size: 1.2em;"> The TetR repressor exists in 2 states: silent and active. In the absence of inducers, the active state of TetR binds to the TetO1 and TetO2 operator sites of the <i>tetp</i> promoter, thus inhibiting transcription. In the presence of inducers, however, TetR undergoes allosteric rearrangement and switches to its silent state, resulting in its release from the promoter, thus allowing transcription to take place.
 
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Revision as of 04:39, 12 October 2016

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tetp Module



The tetp module consists of a Tet repressible promoter upstream of the CDSs of phlf and lacI, which are then followed by the reporter gene, mtagbfp. When the ptet module is active, the two repressor proteins, PhlF and LacI will be expressed along with the reporter gene, mTagBFP. This results in the repression of the two other modules in our system, pphlf and plac.

Promoter, Repressor and Inducer Specification


tetp promoter
  • Promoter name: tetp (Tet repressible promoter)
  • Length: 54 base pairs
  • Strength with:
    • Weak RBS (BBa_B0032): 5.6 - 517.5 (+17) RPU
    • Medium RBS (BBa_B0030): 25.5 - 2323.1 (+259.5) RPU
    • Strong RBS (BBa_B0034): 39.6 - 4041.5 (+247.8) RPU

In gram negative bacteria, TetR represses tetp which regulates tetracycline resistance by driving the expression of TetR and TetA, the latter of which codes for a tetracycline-magnesium complex that pumps tetracycline out of the cell.

  • Repressor name: TetR (Tet repressor)
  • Part Length: 685 base pairs
  • Structure: Homodimer of alpha-helices
  • Binding affinity: To tetp, very light, 0-1 µl/ml has been shown to cause a 5 order of magnitude change in fluorescent protein production

The TetR repressor exists in 2 states: silent and active. In the absence of inducers, the active state of TetR binds to the TetO1 and TetO2 operator sites of the tetp promoter, thus inhibiting transcription. In the presence of inducers, however, TetR undergoes allosteric rearrangement and switches to its silent state, resulting in its release from the promoter, thus allowing transcription to take place.

  • Inducer name: ATc (Anhydrotetracycline)
  • Binding Affinity: (to TetR) Ka ~ 109 M-1
  • Effective range: 250 - 25000 ng/ml

Mechanism of tetp - TetR - ATc Interaction