Part:BBa_K5348005

pL-RBS2

Sequence and Features

Assembly Compatibility:

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

  Illegal BamHI site found at 1874
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 587
  Illegal NgoMIV site found at 659
  Illegal NgoMIV site found at 749
  Illegal NgoMIV site found at 767
  Illegal NgoMIV site found at 1259
  Illegal NgoMIV site found at 1552
  Illegal NgoMIV site found at 1646
  Illegal AgeI site found at 301
  Illegal AgeI site found at 1427
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1316
  Illegal BsaI.rc site found at 200
  

BBa_K5348005 (pL-RBS2)

Construction Design

To reduce the leaky expression of the light-inducible induction system (BBa_K3447133, hereafter referred to as the pL), we reduced the intensity of the RBS linked to the target gene in this element by 108-fold (RBS2). The composition of this element is shown below.

Figure 1. Schematic diagram of the pL-RBS2

Engineering Principle

Under dark condition, histidine kinase (YF1) phosphorylates FixJ (response regulator of histidine kinase), which activates PFixK2 (the target gene for transcription upon FixJ activation), driving the expression of the cI gene (λ phage repressor), which represses the transcription of its cognate promoter, PR (the cognate promoter of cI), and downstream genes cannot be expressed. Under blue light, the cI gene cannot be expressed, PR can be transcribed normally, and downstream genes can be expressed [1].

Reference

[1] H, Mays RL, Hoffman SM, Avalos JL. Optogenetic Control of Microbial Consortia Populations for Chemical Production. ACS Synth Biol. 2021 Aug 20;10(8):2015-2029.