Part:BBa_K1725351

K1725080.luxCDE - bright

Part K1725351 is a luxCDE gene assembly under the R00N1 (BBa_K1725080) promoter generated by BioBrick assembly by combining luxC (BBa_K1725202), luxD (BBa_K1725203) and luxE (BBa_K1725204) ribosome binding site (RBS) libraries together. RBS BBa_K1725307, BBa_K1725302 and BBa_K1725301 are upstream of luxC, luxD and luxE genes, respectively. The assembly has been shown to induce high levels of bioluminescence in E. coli when co-tranformed with pBAD.luxABG-bright (BBa_K1725350). [http://2015.igem.org/Team:Glasgow/Description More information] about the part.

Design of the RBS library for luxCDE

For the construction of the RBS library, a master sequence based on the RBS B0032 was used. 4 nucleotides within the actual ribosome binding site were randomised giving 32 different B0032-derived RBS variants. The predicted efficiency of each RBS library member was estimated using RBS Library Calculator. Originally, RBS Library was designed for every gene in luxABGCDE operon (BBa_K1725352) in order to optimise bioluminescence in E. coli. 32 different BioBrick RBS variants were incorporated upstream of each of the lux genes by PCR, and different libraries were then combined together by BioBrick assembly. This method could potentially produce over 1 billion different variants of the lux operon. Just for the luxCDE assembly, there are over 32000 different variants.

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iGEM logo painted with E. coli co-transformed with BBa_K1725350 and BBa_K1725351

Design of the RBS library for the lux operon

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 1
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 2050
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1435
  Illegal SapI.rc site found at 2603