Part:BBa_K3831034

Construct Z: phosphate-dependent BMC regulation

The composite part Z is composed of certain components of composite parts A (BBa_K3831030), C (BBa_K3831032), and D (BBa_K3831033). Composite part Z will be assembled by modifying composite part D. Promotor Phyperspank would be replaced by the PPho promoter from composite part C which responds to the phosphate concentration in the growth medium. This modification would be done using restriction sites HindIII and NcoI. Additionally, the GFP sequence would be replaced by the sequences of the Pdu BMC shell proteins from composite part A. This modification would be done using restriction sites NdeI and XhoI. By using this clever design, we were able to avoid having to pay for the synthesis of another large construct while making very efficient use of the other constructs.

This part has been combined from previously mentioned parts A, C, and D. Using this concept, we are able to combine our composite parts and test the entire system. Using this part, we will test whether BMC shall proteins are expressed and correctly assembled when under the control of the modified Pgrac Oc promoter. We can also verify the production of the cI repressor via mScarlet-I. At low phosphate concentrations, PPho promoter should activate the transcription of the cI sequence and lead to the production of the cI repressor. This repressor inhibits transcription initiation from the Pgrac Oc promoter and thus prevents the formation of more BMCs. This system ensures that BMCs are produced only when the bacteria encounter high phosphate concentrations.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 4749
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 626
  Illegal AgeI site found at 3891
  
• 1000
  COMPATIBLE WITH RFC[1000]