Part:BBa_K3064026
9xGSP-GFP
This composite part is made up of a kind of improved promoter which is sensitive to particular high blood glucose concentration and EGFP sequence. The introduction of EGFP makes it possible to examine the expression of this composite part. As designed,the part can be used as a glucose-sensing promoter with GFP reporter system.
Usage and Biology
Because there is high blood glucose concentration in diabetics, in order to get improved gene which is sensitive to diabetics it’s convenient to employ promoter which reacts to high blood glucose. Transcription factor -- ChREBP -- can be effectively expressed with high blood glucose. Meanwhile, heterodimer which consists of ChREBP and Mlx can combine with gene promoter CHoRE to induce gene transcription. Therefore, we choose CHoRE as promoter of engineered plasmid and connect it with minP to indicates a transcription start site. In addition, we increase the number of CHoRE to realize that engineered gene will be activated by particular high blood glucose concentration instead of normal blood glucose concentration. Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Characterization
For characterizaton, this part BBa_K3064026 was cloned in pcDNA3.1+ and transfected into HepG2 cell lines using Invitrogen LipofectamineTM 3000. Protocol could be found in our Experiment page. After transfection 12h, we starve the HepG2 cells with glucose-free culture and stimulate with 20mM glucose concentration culture after 6 more hours. Glucose stimulation intensity controlled at 20mM. Sample tested after transfection of 48h.
Figure 1. Firefly luciferase/renilla luciferase RLU for minipromoter and 3/6/9 glucose sensing promoter.
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