Part:BBa_K1592005

GalBD-CRY2 Fusion for Yeast-Two-Hybrid

Cryptochrome 2 (CRY2) is a blue light stimulated photoreceptor, when exposed to blue light, it would interact with CIB1. The CRY2/CIB1 interaction is entirely genetically encoded and does not require addition of any exogenous cofactors. The binding naturally reverses within minutes in the dark, allowing rapid shutoff of transcription by placing samples in the dark. This fusion protein is for use in a yeast-two-hybrid system, and a Gal4 DNA binding domain fused to its C terminus. To regulate DNA transcription by blue light, the system is based on a two-hybrid interaction in which a light-mediated protein interaction brings together two halves (a binding domain and an activation domain) of a split transcription factor. If we remove the stimulation of blue light, dark reversion of CRY2 will dissociate the interaction with CIB1 and halt Gal4-dependent transcription.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 849
  Illegal BglII site found at 1308
  Illegal BamHI site found at 1793
  Illegal XhoI site found at 218
  Illegal XhoI site found at 445
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 733
  Illegal AgeI site found at 1462
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 137
  Illegal BsaI site found at 1085
  Illegal BsaI.rc site found at 494
  Illegal SapI.rc site found at 602