Part:BBa_K3739061

J23100-B0030-blrA-GFP-B0010

blrA can be activated by blue light and produce reactive oxygen species(ROS).

Biology

Flavin-combined fluorescent protein, coded by a kind of optogenetic toolbox LOV(light-oxygen voltage)-based photosensitizer gene, can induce the light-driven killing of bacteria. BlrA, originating from Bacillus subtilis, is a blue light-induced GTP-binding receptor, which possesses the LOV domain and autofluorescence. LOV domain enables it to be activated by blue light and produce ROS, which can damage the bacterial structure.

Gfp codes for a green fluorescence protein.The fluorescence density of GFP can reflect the expression level of BlrA.

Usage

To intuitionally show whether blrA expresses or not, it is linked to a GFP through a 3GS linker. The growth condition and the level of BlrA can be reflected by the intensity of green fluorescence.

Fig.1. The result of regular PCR. Plasmid pET-28a(+).

We take blrA as the core of our kill switch, combine it with promotor, terminator and a series of necessary elements. The whole part is inserted into a plasmid backbone and finally transformed into E.coli to gain enough correct and functional circuits. The circuits are transformed into Vibrio.natriegens. We choose growth curve and CFU to measure the killing effect of BlrA.

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
  INCOMPATIBLE WITH RFC[21]

  Illegal XhoI site found at 470
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 322
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 221
  Illegal SapI site found at 539
  Illegal SapI.rc site found at 902