E colock
From 2006.igem.org
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| - | + | = '''E colock''' ''"self-cycling-E. coli"'' = | |
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[["E colock" Chiba 2006|'''⇒Go to "E colock" English page??'''<br>wikiページ（英語）]] | [["E colock" Chiba 2006|'''⇒Go to "E colock" English page??'''<br>wikiページ（英語）]] | ||
| + | = Concept = | ||
| + | [[Image:Ecolock model.jpg|left|330px]] | ||
| - | + | '''Installing Kai system,<br>E.coli starts oscillating!''' | |
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| + | = Member = | ||
[[Chiba University, Japan 2006|Chiba 2006 wikiページ（英語）]] <br> | [[Chiba University, Japan 2006|Chiba 2006 wikiページ（英語）]] <br> | ||
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*[[User:Matsuno|松野 伸哉]] | *[[User:Matsuno|松野 伸哉]] | ||
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| - | + | = Kai system = | |
| - | + | [[Image:Kai system.jpg|right|400px]]<br /> | |
| - | + | Kai system is generated by 3 proteins(KaiA,KaiB, and KaiC).<br /> | |
| - | + | This is the most simplest circardian oscillation system.<br /> | |
| + | But,it is not known how this oscillation regulate global expression of many proteins in cyanobacteria. | ||
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| + | SasA is a Sensory Kinase(a kind of Histidine Kinase).<br /> | ||
| + | SasA is believed to connect with Kai cycle and transcriptional oscillation of many proteins.<br /> | ||
| + | RpaA is coganate response regulator for SasA, and it's deletion results in the complete loss of transcription oscillation.<br /> | ||
| + | This is all known about Kai. | ||
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| + | '''reference ''' | ||
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| + | [[image:Cyanobavteria.JPG|right|210px]] | ||
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| + | = E.colock Project = | ||
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| + | SasA and RpaA is highly homologous to EnvZ and OmpR,respectively. | ||
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| + | Believing that SasA is communicating with Kai oscillator. | ||
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| + | Believing that RpaA communicate with SasA. | ||
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| + | We designed RpaA-OmpR chimera,in hope that we read our Kai oscillation using ''ompC'' promoter. | ||
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| + | [[Image:Ecolock.jpg|right|400px]]<br /> | ||
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| + | = Chimera design = | ||
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| + | In general, Response Regulator has 2 domeins, | ||
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| + | regulator domein receives phospate from Hiatidine Kinase and effector domein bind to specific sequence of DNA. | ||
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| + | We create 36 chimera of RpaA-OmpR. | ||
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| + | There are different in cutting point, but all of them have recerver domein of RpaA to communicate | ||
| + | SasA,and DNA-binding domein of OmpR to bind and activete right to ''OmpC'' promoter. | ||
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| + | The precedure(↓) | ||
| + | We used 6 excite primers on each RpaA and OmpR， | ||
| + | It can make 6 ×6 = 36 different chimeras!! | ||
| + | [[Image:Excite PCR.JPG|600px]] | ||
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| - | + | = Next step (＊_＊)/ = | |
| - | + | The construct of chimeras is finished! | |
| - | + | Chimaras along with WT RpaA and OmpR will be tested in their function as response regulator that switch ON/OFF the ''ompC''promoter. | |
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| + | [[Image:Jikkensitai.JPG|600px]] | ||
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| - | + | We will find some chimeras indeed pick up Kai cycle. | |
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| - | = | + | = Last up date = |
| - | + | *11/15 | |
Revision as of 13:34, 15 November 2006
Contents |
E colock "self-cycling-E. coli"
⇒Go to "E colock" English page??
wikiページ（英語）
Concept
Installing Kai system,
E.coli starts oscillating!
Member
Chiba 2006 wikiページ（英語）
[http://chem.tf.chiba-u.jp/igem/ iGEM Chiba オフィシャルページ（日本語）]
『む』チーム
Kai system
Kai system is generated by 3 proteins(KaiA,KaiB, and KaiC).
This is the most simplest circardian oscillation system.
But,it is not known how this oscillation regulate global expression of many proteins in cyanobacteria.
SasA is a Sensory Kinase(a kind of Histidine Kinase).
SasA is believed to connect with Kai cycle and transcriptional oscillation of many proteins.
RpaA is coganate response regulator for SasA, and it's deletion results in the complete loss of transcription oscillation.
This is all known about Kai.
reference
E.colock Project
SasA and RpaA is highly homologous to EnvZ and OmpR,respectively.
Believing that SasA is communicating with Kai oscillator.
Believing that RpaA communicate with SasA.
We designed RpaA-OmpR chimera,in hope that we read our Kai oscillation using ompC promoter.
Chimera design
In general, Response Regulator has 2 domeins,
regulator domein receives phospate from Hiatidine Kinase and effector domein bind to specific sequence of DNA.
We create 36 chimera of RpaA-OmpR.
There are different in cutting point, but all of them have recerver domein of RpaA to communicate SasA,and DNA-binding domein of OmpR to bind and activete right to OmpC promoter.
The precedure(↓)
We used 6 excite primers on each RpaA and OmpR，
It can make 6 ×6 = 36 different chimeras!!
Next step (＊_＊)/
The construct of chimeras is finished! Chimaras along with WT RpaA and OmpR will be tested in their function as response regulator that switch ON/OFF the ompCpromoter.
We will find some chimeras indeed pick up Kai cycle.
Last up date
- 11/15
