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Part:BBa_K2100032:Experience

Designed by: Kathleen Brandes   Group: iGEM16_MIT   (2016-10-17)
Revision as of 19:18, 21 October 2016 by Kbrandes (Talk | contribs) (→‎Applications of BBa_K2100032)


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Applications of BBa_K2100032

We used our promoter to build the pHybrid:eYFP construct to characterize the functionality of our promoter.

We characterized the synthetic pHybrid promoter in two cell lines: MCF7 and tHESC. All cell lines have endogeneous progesterone receptor A as well as estrogen receptor alpha. We analyzed data from cells induced with 1 uM of MPA compared to those uninduced with hormones. This concentration of MPA was recommended to us as an appropriate "on" or saturating concentration by the Griffith Lab which provided us with these cells.

We transfected tHESC cells with 250ng of hEF1a:mKate as a transfection marker and 250 ng pPRE4:eYFP to examine the promoter's transcriptional activity by observing increases in yellow fluorescence upon cells being induced with 1 uM MPA. This ratio was chosen to be 1:1 based on the small amount of plasmids being transfected. We were looking to deterimne the on-off functionality of our promoter pPRE4 in the tHESC cell line.

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UNIQ7094d75ea9fd3872-partinfo-00000000-QINU UNIQ7094d75ea9fd3872-partinfo-00000001-QINU