Part:BBa_K1119008

CMV promoter - GFP - hGH polyA tail

In characterization of Mitochondrial Leader Sequence (BBa_K1119001), negative control was made by GFP generator that does not contains the CDS of MLS (BBa_K1119008). The translation unit was driven by CMV promoter (BBa_K1119006) and terminated by hGH polyA signal (BBa_K404108).

Result

In the characterization of CMV promoter (BBa_K1119006) , the coding sequence of CMV promoter was assembled with GFP reporter (BBa_K648013) and hGH polyA terminator (BBa_K404108) using Freiburg’s RFC25 format.

The pCMV-GFP was then transfected into HEK293FT cells and in vivo green fluorescence signal was observed under confocal microscope.

The positive control was pEGFP-N1 (Clontech) that contains CMV promoter and EGFP reporter. A negative control was made by GFP generator that does not contain the CMV promoter.

The [http://2013.igem.org/Team:Hong_Kong_HKUST/characterization detailed protocol] of our characterization can be found in HKUST iGEM 2013 Wiki.

Figure 1. CMV promoter drives expression of GFP. HEK cells transfected with pCMV-GFP expressed GFP Signal. HEK cells transfected with the commercial pEGFP-N1 showed similar results, while the negative control GFP without any promoter was not expressed.

Sequence and Features

Assembly Compatibility:

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

  Illegal BglII site found at 614
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 628
  Illegal AgeI site found at 1369
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1772
  Illegal BsaI.rc site found at 1274