Difference between revisions of "Part:BBa K3064012"
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===Usage and Biology=== | ===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 | + | 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 transcription¹.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. |
| − | In this composite part, we used nine ChoRE and | + | In this composite part, we used one minP and nine ChoRE and apply luciferase gene as downstream reporter in plasmid PGL3-9XGSP to verify its ability. |
| + | |||
| + | ===Materials=== | ||
| + | |||
| + | PGL3-9XGSP | ||
| + | |||
| + | PGL3-minP | ||
| + | |||
| + | Hepg2 cell line | ||
| + | |||
| + | Dual Luciferase Reporter Gene Assay Kit from from Beyotime company | ||
| + | |||
| + | ===Method=== | ||
| + | |||
| + | We separately transfected plasmids PGL3-9XGSP and into hepg2 cells. After 48 hours, these cells were gathered and dissoved for luciferase expression test with Dual Luciferase Reporter Gene Assay Kit of Beyotime company. | ||
| + | |||
| + | ===Result=== | ||
| + | |||
| + | Taking renilla as the internal reference and comparing with the control experiment, it’s obvious to find that CHoRE can significantly improve minP promoter’s promoting intensity. | ||
| + | https://static.igem.org/mediawiki/parts/9/9a/T--NUDT_CHINA--2019pic9xgsp.png | ||
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Revision as of 23:30, 20 October 2019
9XGlucose Sensing Promoter
This composite part is a kind of improved promoter which is sensitive to particular high blood glucose concentration.
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 transcription¹.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.
In this composite part, we used one minP and nine ChoRE and apply luciferase gene as downstream reporter in plasmid PGL3-9XGSP to verify its ability.
Materials
PGL3-9XGSP
PGL3-minP
Hepg2 cell line
Dual Luciferase Reporter Gene Assay Kit from from Beyotime company
Method
We separately transfected plasmids PGL3-9XGSP and into hepg2 cells. After 48 hours, these cells were gathered and dissoved for luciferase expression test with Dual Luciferase Reporter Gene Assay Kit of Beyotime company.
Result
Taking renilla as the internal reference and comparing with the control experiment, it’s obvious to find that CHoRE can significantly improve minP promoter’s promoting intensity.
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
