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Part:BBa_K1119008

Designed by: Hyun Jung Lee   Group: iGEM13_Hong_Kong_HKUST   (2013-09-16)
Revision as of 17:07, 30 October 2017 by Danakd (Talk | contribs) (Improved Part)

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CMV promoter - GFP - hGH polyA tail

This is a mammalian GFP generator used for characterization of our CMV promoter (BBa_K1119006). This GFP generator contains the CMV promoter sequence with GFP reporter in RFC25 format(BBa_K648013) and hGH polyA terminator (BBa_K404108).

This GFP generator was then transfected into HEK293FT cells and in vivo green fluorescence signal was observed under confocal microscope.

pEGFP-N1 (Clontech) served as the positive control. It contains the CMV promoter and EGFP reporter. A GFP generator that does not contain the CMV promoter was used as the negative control.

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 gave GFP signals. HEK cells transfected with the commercial pEGFP-N1 showed similar results, while the same construct without any promoter did not give any GFP signals.

Improved Part

iGEM Technion 2017 improved this part to BBa_K2520006. This improved device contains a TRE promoter instead of the original CMV, to allow for inducible expression of GFP within mammalian cells.


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


[edit]
Categories
//chassis/eukaryote/human
Parameters
None