DNA

Part:BBa_K2100003

Designed by: Kathleen Brandes   Group: iGEM16_MIT   (2016-10-10)


pENTR hEF1a

The pENTR hEF1a part is an entry vector containing the human elongation factor 1 alpha (hEF1a) promoter, a constituitive mammalian promoter.

We have characterized the hEF1a promoter in all four cell lines we experimented on in the duration of our project using the following fluorescent tags: eYFP, mKate, and tagBFP.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 6
    Illegal EcoRI site found at 1210
    Illegal PstI site found at 337
    Illegal PstI site found at 842
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 6
    Illegal EcoRI site found at 1210
    Illegal PstI site found at 337
    Illegal PstI site found at 842
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 6
    Illegal EcoRI site found at 1210
    Illegal BglII site found at 591
    Illegal BamHI site found at 1197
    Illegal XhoI site found at 990
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 6
    Illegal EcoRI site found at 1210
    Illegal PstI site found at 337
    Illegal PstI site found at 842
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 6
    Illegal EcoRI site found at 1210
    Illegal PstI site found at 337
    Illegal PstI site found at 842
    Illegal NgoMIV site found at 725
    Illegal AgeI site found at 103
  • 1000
    COMPATIBLE WITH RFC[1000]

The MIT iGEM team used the hEF1a promoter to constitutively express fluorescent proteins as transfection markers in our experiments in HEK293, tHESC, MCF-7, and ISH cells. The amount of fluorescent units observed is an indication of the number of plasmids a particular cell uptakes. The graph below shows two single color controls from one of our experiments. We transfected one well of HEK293 cells with hEF1a-mKate and another with hEF1a-eYFP. The cells that uptook the hEF1a-mKate plasmids showed an increase in only the red fluorescent output, while the cells that uptook the hEF1a-eYFP plasmids showed a clear increase in only the yellow fluorescent output.

T--MIT--hEF1a.png



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Categories
//awards/part_collection/2016
Parameters
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