Part:BBa_K2865008
BNP promoter-eGFP-poly(A)
This is a mammalian eGFP generator used for characterization of our BNP promoter (BBa_K2865000). This eGFP generator contains the BNP promoter sequence followed by eGFP reporter in RFC25 format(BBa_K2865005) and SV40 polyA terminator (BBa_K2865004). This eGFP generator was then transfected into H9C2 cells and in vivo green fluorescence signal was observed under fluorescence microscope.
Experiment
Fluorescence microscope
We transfected H9C2 cell with this part by liposome transfection. And we select two widely used chemicals to simulate cell microenvironment in heart failure, ET1 and Ang II. From scientists’ previous work, we learnt that the appropriate concentration is 10-7mol/L and BNP is in really high expression in 10-7mol/L.
After 48h of expression, we detected all the cells using Fluorescence microscope. However, the BNP promoter did not work very well. All the groups using ET1 and AngII have no green fluorescent signal. At the same time, the groups using CMV promoter succeeded in expressing eGFP.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 1284
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 242
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