Part:BBa_K2587027

luxI_luxR_P_lux_gfp

This construct comprises parts from the Quorum sensing system from Vibrio fischeri. It contains the LuxI, the acyl homoserine lactone synthase, the LuxR, the regulator, which will bind to the promoter pLux, that will induce synthesis of GFP.

Usage and Biology

• Acyl homoserine lactone synthase: LuxI
• Regulator: LuxR
• Promoter: pLux, inducible by the quorum sensing molecule acyl homoserine lactone bound to LuxR
• GFPfluorescent protein
• GFP expression is induced after synthesis of the acyl homerine lactone by LuxI, which binds with LuxR to the pLux promoter
• This construct shows functionality of promoter by increase of fluorescence upon time
• This part contains Type II S restriction sites and can be further used for scarless assembly with other parts

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
  Illegal NheI site found at 829
  Illegal NheI site found at 852
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1082
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 892
  Illegal BsaI site found at 1681
  Illegal BsaI.rc site found at 1523
  Illegal BsaI.rc site found at 1790
  Illegal BsaI.rc site found at 2468
  

Characterization

Characterisation of promoter by fluorescence measurement of GFP
To check whether the promoter, pLux, is really induced by the binding of the acyl homoserine lactone to the LuxR, a further construct (BBa_K2587027) is created which has the lysis gene E exchanged by a GFP gene. Here the relative fluorescence units are measured in comparison to the wild type E.coli cells.

Results It can clearly be observed, that fluorescence increases upon growth of the cells. Wild type cells, as a negative control, show little or no fluorescence at all.

Figure 1: Relative fluorescence measurement of lysis construct and E.coli wild type cells (negative control). Exctinction: 485nm; Emission: 520nm. Cells were measured only within a time span of 24h.