Part:BBa_K1896016

INP_RC-mGFPuv2 generator

This part contains the INP_RC-mGFPuv2 fusion protein controlled by a weak to medium constitutive promoter and RBS.

Usage and Biology

INP_RC is a truncated Ice Nucleating Protein from which the N-terminal domain has been removed. The truncated INP is still capable of catalysing the formation of ice crystals, but is no longer localised to the outer cell membrane [1]. A monomeric GFP variant (mGFPuv2) has been fused to INP_RC to act as a control in protein adhesion experiments performed by the UGent Belgium 2016 iGEM team.

When crude cell lysate of a culture of E. coli expressing mGFPuv2 is added to supercooled water, no reaction takes place. When crude cell lysate containing INP_RC-mGFPuv2 is added, ice is formed instantly. This demonstrates that the cytoplasmatic INP_RC fusion protein is still active.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 203
  Illegal NgoMIV site found at 1715
  Illegal NgoMIV site found at 1859
  Illegal NgoMIV site found at 2123
  Illegal NgoMIV site found at 2267
  Illegal NgoMIV site found at 2315
  Illegal NgoMIV site found at 2435
  Illegal AgeI site found at 1379
  
• 1000
  COMPATIBLE WITH RFC[1000]

References

1. Green, R. L., Corotto, L. V., & Warren, G. J. (1988). Deletion mutagenesis of the ice nucleation gene from Pseudomonas syringae S203. Molecular and General Genetics MGG, 215(1), 165-172.