Part:BBa_K2957050

CMV-2xGal4 (Gal4 repressible promoter), iRFP720

Contents:CMV-2xGal4 (Gal4 repressible promoter), iRFP720, SynthpA, with Carb resistance

What is it?: A Composite part; it is a genetic circuit with CMV-2xGal4 promoter and iRFP720 gene allowing for inducible expression of the reporter iRFP720.

What does it do?: This part has a promoter that is only active when induced by the syn-notch receptor and was put in our genetic circuits to show inducible production of the iRFP720. It was important for our project in particular because it was able to show successful transfection in HEK cells and demonstrate inducible expression in the HEK cells. This specific part was important to show a wave mechanic since when neutrophils expressing the syn-notch antigen would bind to the receptor on HEK cells, the promoter can activate more or less iRFP720 production.

How to use it?: This part was created with Type IIS Cloning and transfected into HEKs. We then were able to test for expression of iRFP720. See more on the Parts Overview Page or MIT iGEM 2019 wiki: <a href=https://2019.igem.org/Team:MIT/Parts>.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal XbaI site found at 1630
  Illegal SpeI site found at 92
  Illegal PstI site found at 1091
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal SpeI site found at 92
  Illegal PstI site found at 1091
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 677
  Illegal XhoI site found at 1013
  Illegal XhoI site found at 1019
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal XbaI site found at 1630
  Illegal SpeI site found at 92
  Illegal PstI site found at 1091
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal XbaI site found at 1630
  Illegal SpeI site found at 92
  Illegal PstI site found at 1091
  Illegal NgoMIV site found at 794
  Illegal AgeI site found at 1636
  
• 1000
  COMPATIBLE WITH RFC[1000]