Part:BBa_K2638560

New designed control vector with eCFP for the measurement of the expression strength

In synthetic biology the control of transcription and translation is of enormous importance. Therefore, promoters and ribosome binding sites (RBS) play a central role in each iGEM project. The selection of a suitable promoter/RBS combination is absolutely necessary to achieve the appropriate expression level for the respective purpose. We tested different combinations of known promoters and RBS, which are often used by the iGEM community. To test the expression strength of these promoter-RBS combinations we constructed a modified pSB1C3 backbone. An eCFP (Part:BBa_E0022) under the control of a promoter from the Anderson collection, a synthetic RBS and two terminators were integrated into the pSB1C3 backbone. This expression unit is used as a reference. To test the expression strength of other promoter-RBS combinations a second reporter gene mRFP (Part:BBa_E1010) under control of the respective target promoter-RBS combination was integrated between the pSB1C3 prefix and suffix. With the combination of the reference reporter gene in the backbone and the mRFP integrated between the pSB1C3 prefix and suffix, the expression strength of the different promoter-RBS combinations can be normalized to the certain expression of the reporter gene in the backbone. The detection of the differential expression strength of the different promoter-RBS combinations can be calculated through the ration of the fluorescence signals at 608 nm and 485 nm. For the detailed characterization see Part:BBa_K2638560.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal prefix found in sequence at 2973
  Illegal suffix found in sequence at 1
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 2973
  Illegal NheI site found at 896
  Illegal NheI site found at 919
  Illegal SpeI site found at 2
  Illegal PstI site found at 16
  Illegal NotI site found at 9
  Illegal NotI site found at 2979
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 2973
  Illegal XhoI site found at 1957
  Illegal XhoI site found at 2849
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal prefix found in sequence at 2973
  Illegal suffix found in sequence at 2
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found in sequence at 2973
  Illegal XbaI site found at 2988
  Illegal SpeI site found at 2
  Illegal PstI site found at 16
  
• 1000
  COMPATIBLE WITH RFC[1000]