Team:Fudan-Shanghai/Project

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<td rowspan=2 bgcolor=#FFFFCE width=400  ><h1>Detail:</h1><font size=3>
<td rowspan=2 bgcolor=#FFFFCE width=400  ><h1>Detail:</h1><font size=3>
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The project “E.tree” is consisted of two genetically modified E.coli: E. trunk and E.leaf. <br><br>
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<b>E.trunk bacteria</b> detect nitrates in the medium and release either LasI or RhlI directed signals. It includes two circuits: the first pathway is controlled by a nitrate sensitive promoter (pYeaR), so that if the medium contains nitrates, the entire pathway will be on, RhlI (produces AHL C4-homoserine lactone), tetR and GFP will be produced. TetR then binds with pTet in the second pathway and blocks it. <br>
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E.trunk detects nitrates in the medium; if KNO3 is present, the first circuit is activated, RhlI produces the AHL (C4-homoserine lactone) and TetR is synthesized to block the second pathway. If the medium lacks KNO3, the first pathway is off while the second pathway is activated and LasI produces an AHL autoinducer (3OC12-homoserine lactone). <br><br>
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If the medium contains no nitrates, the first pathway is repressed while the second one is unblocked, so LasI (produces 3OC12-homoserine lactone) and RFP will be synthesized. <br>
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E.leaf changes color corresponding to the signal molecules released by E.trunk. RhlR synthesized in E.leaf binds to the RhlI-directed signal, and the complex then interact with the pRhl promoter and yellow pigment is produced. This circuit also encodes RhlI, which multiple the circuit, so that all E.leaf would turn yellow. Similarly, if E.trunk releases 3OC12-homoserine lactone, which is encodes by LasI, all “leaves” would turn green. <br><br>
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<b>E.leaf bacteria </b>change color according to the signal molecules released by E.trunk. LasR and RhlR are constitutive synthesized in E.leaf; they bind to the LasI and RhlI directed signals, respectively. <br>
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<br><b>Overall effect:</b>
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  If the medium contains nitrates, E.trunk releases C4-homoserine lactone (the product of RhlI); the signal molecules enter E.leaf bacteria and bind to RhlR. This RhlI/R complex then interact with pRhl promoter and green fluorescence is emitted. Meanwhile, RhlI molecules further synthesized in E.leaf bacteria, so that not only bacteria grow along the border but all E.leaf bacteria on the plate will turn green. <br>
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  Similarly, if the medium lacks nitrates, E.trunk bacteria release 3OC12-homoserine lactone (the products of LasI) and the “leaves” turn yellow.
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<td bgcolor=#CEFFCE align=center><a href="https://static.igem.org/mediawiki/2011/a/a9/E.tree_detail.JPG"><img src="https://static.igem.org/mediawiki/2011/a/a9/E.tree_detail.JPG" height=300></a></td>
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<td><a href="https://static.igem.org/mediawiki/2011/e/e8/Fudan_Testall.JPG"><img src="https://static.igem.org/mediawiki/2011/a/aa/Fudan_exam.JPG" width=500></a></td>
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<td colspan=2><h1><img src="https://static.igem.org/mediawiki/2011/e/ee/Fudan_Etree.jpg " alt="E.tree" width=100  align=center hspace=20><b> Part II: Dinner Service </b></h1></td>
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<td colspan=2><h1><img src="https://static.igem.org/mediawiki/2011/9/96/DINER_SERVICE.jpg" alt="E.DinnerService" width=100  align=center hspace=20><b> Part II: Dinner Service </b></h1></td>
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<td rowspan=2 bgcolor=#FFE6D9 width=400  ><h1>Detail:</h1><font size=3>
<td rowspan=2 bgcolor=#FFE6D9 width=400  ><h1>Detail:</h1><font size=3>
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Our project was designed to show the quorum sensing system between bacterias in the background story of "dinner service". To construct our pathway, we chose J23112 as promoter, using AHL and aiiA as the sensing materials  with the reporter of RFP and YFP.<br><br>
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<b>The "customer" E.coli </b>is consisted of two pathways.
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Two circumstances were constructed to represent the"customer" and the "chef".The hungry "customer" express luxI to catalyze the synthesis of AHL, with the red fluorescent as a signal. AHL binds to the luxr part of the"chef" and stimulates the right hand lux promoter. The part BBa_K091109 works to generate luxS, a synthase that produces DPD and forms the AI-2 signaling molecule and express YFP to show yellow fluorescent.<br><br>
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The first pathway codes IsrR, a siganl that activates the second pathway, and aiiA, which degrades IsrR. The promoter for IsrR is stronger than that of aiiA. In that case, part of the IsrR will be degraded, but still some will be able to activate the second pathway. When the second pathway is activated, LuxI is produced and CFP emitted.<br>
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With the stimulating signal of "chef", which means the "dinner", the "customer " is satisfied and the promoter J23112 starts to express aiia to inhibite the AHL signal and "chef" response. The fluorescent disappears and the process is stoped
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<b>The "chef" E.coli</b> also includes two pathways. The first synthesize LuxR, in the presence of AHL( the product of LuxI), it binds to the promoter LuxR and LuxS( the meal) is served, YFP is emited to show this process.<br><br>
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In sum, the hungry customer orders the meal by giving out signal "luxI". The "chef" containing luxR, senses the "order" signal and prepares for the meal-LuxS. The dinner service is accompanied with yellow fluorescent.
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When the customer is full, IsrR synthesis is inhibited, the aiiA degrades the remaining IsrR in the cell, thus the production of LuxI is stopped. And meal is no longer served.
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<td bgcolor=#E2C2DE align=center><a href="https://static.igem.org/mediawiki/2011/a/a4/DinnerService.JPG"><img src="https://static.igem.org/mediawiki/2011/a/a4/DinnerService.JPG" height=300 alt="dinner service"></a></td>
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<td colspan=2><h1><img src="https://static.igem.org/mediawiki/2011/e/ee/Fudan_Etree.jpg " alt="E.tree" width=100 align=center hspace=20><b> Part III: Neon Lights </b></h1></td>
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<td colspan=2><h1><img src="https://static.igem.org/mediawiki/2011/1/11/Fudan_neonLights.jpg" alt="E.Lights" width=100 align=center hspace=20><b> Part III: Neon Lights </b></h1></td>
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<td  bgcolor= #ECECFF width=600  ><h1>Detail:</h1><font size=3>
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<td  bgcolor= #FFECF5 width=600  ><h1>Detail:</h1><font size=3>
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In the project of neon lights, we intended to have E.coli engineered for emitting one color of fluorescence during exponential phase and another color during stationary phase, so that it is convenient to identify the phase and may also be utilized for other purposes. To achieve this goal, we chose BBa_J45992, a stationary phase promoter, and BBa_J45996, only active during exponential phase. Then we tried to produce different luciferases under the control of the promoters. Parts of various luciferases have been obtained by the team of Cambridge in 2010, for example, BBa_K325210, BBa_K325229, etc. System of degradation, such as ClpXP, can be used to optimize the project by degrading the previous luciferase when the latter is being produced, to make the latter color purer and the process of color changing faster. In addition, the fluctuation of the lusiferase activity due to the growth should be taken into consideration.
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In the project of neon lights, we intended to have E.coli engineered for emitting one color of luminescence during exponential phase and another color during stationary phase, so that it is convenient to identify the growth phase and may also be utilized for other purposes.<br>
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To achieve this goal, we chose BBa_J45992, a stationary phase promoter, and BBa_J45996, a promoter that is only active during exponential phase and assemble different luciferase under the control of the promoters. Parts of various luciferases have been obtained from the registry submitted by the team of Cambridge in 2010, for example, BBa_K325210, BBa_K325229, etc. System of degradation, such as ClpXP, can be used to optimize the project by degrading the previous luciferase when the latter is being produced, to make the latter color purer and the process of color changing faster. In addition, the fluctuation of the lusiferase activity due to the growth should be taken into consideration.  
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<td bgcolor=#FFFFCE align=center><a href="https://static.igem.org/mediawiki/2011/d/d1/Neonlights.JPG"><img src="https://static.igem.org/mediawiki/2011/d/d1/Neonlights.JPG" width=300></a></td>
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<td bgcolor=#F1E1FF align=center><a href="https://static.igem.org/mediawiki/2011/d/d1/Neonlights.JPG"><img src="https://static.igem.org/mediawiki/2011/d/d1/Neonlights.JPG" width=300></a></td>
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Latest revision as of 19:50, 5 October 2011


2011 iGEM Fudan-Shanghai Template

2011 iGEM Fudan_Project


E.tree Part I: E.Tree

Detail:

E.trunk bacteria detect nitrates in the medium and release either LasI or RhlI directed signals. It includes two circuits: the first pathway is controlled by a nitrate sensitive promoter (pYeaR), so that if the medium contains nitrates, the entire pathway will be on, RhlI (produces AHL C4-homoserine lactone), tetR and GFP will be produced. TetR then binds with pTet in the second pathway and blocks it.
If the medium contains no nitrates, the first pathway is repressed while the second one is unblocked, so LasI (produces 3OC12-homoserine lactone) and RFP will be synthesized.
E.leaf bacteria change color according to the signal molecules released by E.trunk. LasR and RhlR are constitutive synthesized in E.leaf; they bind to the LasI and RhlI directed signals, respectively.

Overall effect: If the medium contains nitrates, E.trunk releases C4-homoserine lactone (the product of RhlI); the signal molecules enter E.leaf bacteria and bind to RhlR. This RhlI/R complex then interact with pRhl promoter and green fluorescence is emitted. Meanwhile, RhlI molecules further synthesized in E.leaf bacteria, so that not only bacteria grow along the border but all E.leaf bacteria on the plate will turn green.
Similarly, if the medium lacks nitrates, E.trunk bacteria release 3OC12-homoserine lactone (the products of LasI) and the “leaves” turn yellow.

E.DinnerService Part II: Dinner Service

Detail:

The "customer" E.coli is consisted of two pathways. The first pathway codes IsrR, a siganl that activates the second pathway, and aiiA, which degrades IsrR. The promoter for IsrR is stronger than that of aiiA. In that case, part of the IsrR will be degraded, but still some will be able to activate the second pathway. When the second pathway is activated, LuxI is produced and CFP emitted.
The "chef" E.coli also includes two pathways. The first synthesize LuxR, in the presence of AHL( the product of LuxI), it binds to the promoter LuxR and LuxS( the meal) is served, YFP is emited to show this process.

In sum, the hungry customer orders the meal by giving out signal "luxI". The "chef" containing luxR, senses the "order" signal and prepares for the meal-LuxS. The dinner service is accompanied with yellow fluorescent. When the customer is full, IsrR synthesis is inhibited, the aiiA degrades the remaining IsrR in the cell, thus the production of LuxI is stopped. And meal is no longer served. 		dinner service

E.Lights Part III: Neon Lights

Detail:

In the project of neon lights, we intended to have E.coli engineered for emitting one color of luminescence during exponential phase and another color during stationary phase, so that it is convenient to identify the growth phase and may also be utilized for other purposes.
To achieve this goal, we chose BBa_J45992, a stationary phase promoter, and BBa_J45996, a promoter that is only active during exponential phase and assemble different luciferase under the control of the promoters. Parts of various luciferases have been obtained from the registry submitted by the team of Cambridge in 2010, for example, BBa_K325210, BBa_K325229, etc. System of degradation, such as ClpXP, can be used to optimize the project by degrading the previous luciferase when the latter is being produced, to make the latter color purer and the process of color changing faster. In addition, the fluctuation of the lusiferase activity due to the growth should be taken into consideration.

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