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Human DKK1 Promoter
The human DKK1 promoter is the promoter regulating the human DKK1 gene. The promoter is located on human chromosome 10, and will be activated upon DKK1 gene activation.
We used PCR technique to amplify a segment of human chromosome 10 (from 52311944 to 52314294). The estimated length of our DKK1 promoter segment is around 2356bp.
To test the functionality of the DKK1 promoter, we transfected HEK 293 cells with a plasmid (BBa_K2751024) containing the DKK1 promoter and a mCherry reporter protein. We added various concentrations of testosterone to stimulate the DKK1 promoter. The expression of mCherry was read by IX83 confocal microscope using the same exposure time, and we selected a field full of cells from each condition group to control the number of cells observed. Quantification of cells expressing the mCherry protein was determined via Metamorph multi-wavelength cell scoring. Intensity 10 gray levels above local background were determined as real fluorescence emission.
|Picture taken under 10x||Concentration of Testosterone||Number of cells expressing mCherry protein determined by Metamorph|
As the concentration of testosterone rises from 0.05ug/mL to 5ug/mL, the number of cells that express mCherry protein also increases. This shows that testosterone level within a range can stimulate the activation of DKK1 promoter, resulting in increasing expression of mCherry protein.