Difference between revisions of "Team:Toulouse/Notebook"

 
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  <ul>
 
  <ul>
 
         <li><a href="#part1"> January </i></a></li>
 
         <li><a href="#part1"> January </i></a></li>
−
         <li><a href="#part2">  From february to may </a></li>
+
         <li><a href="#part2">  From February to May </a></li>
 
         <li><a href="#part3">  May </a></li>
 
         <li><a href="#part3">  May </a></li>
 
         <li><a href="#part4">  June</a></li>
 
         <li><a href="#part4">  June</a></li>
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−
 
+
 
   <div class="subtitle" id="part1">
 
   <div class="subtitle" id="part1">
 
  <h3>January</h3>
 
  <h3>January</h3>
−
  </div>
+
  </div>  
−
+
<div class="group">
−
<div class="group center">
+
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
 
During this period, we were informed by iGEM toulouse instructors of what this competition is, and what was expected of us.
 
During this period, we were informed by iGEM toulouse instructors of what this competition is, and what was expected of us.
 
  </p>
 
  </p>
 
</div>
 
</div>
 +
 +
 
 +
 +
 
   
 
   
 
    
 
    
 
    
 
    
 
  <div class="subtitle" id="part2">
 
  <div class="subtitle" id="part2">
−
  <h3>From february to may</h3>
+
  <h3>From February to May</h3>
 
  </div>   
 
  </div>   
 
    
 
    
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  <ul style="font-size:15px;">
 
  <ul style="font-size:15px;">
 
  <li>Previous subjects. What are the themes explored by iGEM? Give examples of good projects and explain why they won?</li>
 
  <li>Previous subjects. What are the themes explored by iGEM? Give examples of good projects and explain why they won?</li>
−
  <li>Biobricks? What are they? What are their advantages? What are the materials send by iGEM? How does the registry work?</li>
+
  <li>Biobricks? What are they? What are their advantages? What are the materials sent by iGEM? How does the registry work?</li>
 
  <li>Competition and deadlines. How does the competition works and what are the important deadlines?</li>
 
  <li>Competition and deadlines. How does the competition works and what are the important deadlines?</li>
 
  </ul>
 
  </ul>
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   <div class="group center">
 
   <div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
  We also created our mail address and a google drive to organise our files online.
+
  We also created our email address and a google drive to organise our files online.
 
  </p>
 
  </p>
 
</div>
 
</div>
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  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
 
  <br>
 
  <br>
−
  Every group of two or three students presented one of the subjects seen above. Every presentation was followed by a Q&A so that everyone understood clearly the topic.
+
  Every group of two or three students presented one of the subjects seen above. Every presentation was followed by a Q&A. Thanks to it, everyone understood clearly the topic.
 
  </p>
 
  </p>
 
</div>
 
</div>
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  <li>Strategy</li>
 
  <li>Strategy</li>
 
  <li>Difficulty</li>
 
  <li>Difficulty</li>
−
  <li>Modeling</li>
+
  <li>Modelling</li>
 
  <li>Device</li>
 
  <li>Device</li>
 
  <li>Benefit</li>
 
  <li>Benefit</li>
−
  <li>Ethic</li>
+
  <li>Ethics</li>
 
  <li>Planning</li>
 
  <li>Planning</li>
 
  <li>Why is this project going to win?</li>
 
  <li>Why is this project going to win?</li>
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  <div class="group center">
 
  <div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
The following weeks were spent brainstorming. Everyone presented at least two project ideas. Here are a few examples of projects reviewed:
+
The following weeks were spent brainstorming. Everyone presented at least two project ideas. Here are few examples of projects reviewed:
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="three_quarter">
 
<div class="three_quarter">
 
  <ul style="font-size:15px;">
 
  <ul style="font-size:15px;">
−
  <li>Creation of a rapid detection kit for hepatitis</li>
+
  <li>Creation of a quick detection kit of hepatitis</li>
 
  <li>Degradation of Lignin</li>
 
  <li>Degradation of Lignin</li>
 
  <li>Use of ice nucleation proteins for water desalination</li>
 
  <li>Use of ice nucleation proteins for water desalination</li>
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  Two projects were still in course to be chosen, the use of ice nucleating proteins to purify water and the creation of a  
 
  Two projects were still in course to be chosen, the use of ice nucleating proteins to purify water and the creation of a  
 
  bio trap against the parasitic mite <i>Varroa destructor</i>. This last one was chosen to be our project on the grounds that it  
 
  bio trap against the parasitic mite <i>Varroa destructor</i>. This last one was chosen to be our project on the grounds that it  
−
  was the most feasible. Based on bibliography, the initial idea was to build an engineered bacteria which will have to produce butyrate to attract the parasite. The following weeks were the subject of extensive research on the topic.  
+
  was the most feasible. Based on bibliography, the initial idea was to build an engineered bacteria which will have to produce butyrate to attract the parasite. The following weeks were the subject of extensive researches on the topic.  
 
<br /> <br /> Moreover, it was time to choose attributions to make the project management easier, below in the table, the main assignments. </p>
 
<br /> <br /> Moreover, it was time to choose attributions to make the project management easier, below in the table, the main assignments. </p>
 
  </div>
 
  </div>
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<tr>
 
<tr>
 
<th>Tanchon Melany</th>
 
<th>Tanchon Melany</th>
−
<td>Modeling</td>
+
<td>Modelling</td>
 
   </tr>
 
   </tr>
 
    
 
    
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   <tr>
 
   <tr>
 
<th>Pons Marine</th>
 
<th>Pons Marine</th>
−
<td>Experiences</td>
+
<td>Experiments</td>
 
   </tr>
 
   </tr>
 
    
 
    
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   <tr>
 
   <tr>
 
<th>Chaumont Laetitia</th>
 
<th>Chaumont Laetitia</th>
−
<td>Ethic</td>
+
<td>Ethics</td>
 
   </tr>
 
   </tr>
 
      
 
      
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<div class="group center">
 
<div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
  Two members of our team participated in the Toulouse Exposcience. They presented a poster on synthetic biology and the iGEM competition. They also organized banana DNA extraction experiences for children. This successful event was a good opportunity for us to start learning how to design a poster and to communicate about synthetic biology.
+
  Two members of our team participated in the Toulouse Exposciences. They presented a poster on synthetic biology and the iGEM competition. They also organized banana DNA extraction experiences for children. This successful event was a good opportunity for us to start learning how to design a poster and to communicate about synthetic biology.
 
  </p>
 
  </p>
 
   </div>
 
   </div>
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  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
 
We held our first meeting without instructors to discuss the design of the constructions needed to create the engineered bacteria.  
 
We held our first meeting without instructors to discuss the design of the constructions needed to create the engineered bacteria.  
−
Our reflexion led us to the conclusion that our construction would depend on the bee life cycle. We therefore started searching for beekeepers  
+
Our reflection led us to the conclusion that our construction should respect the bee life cycle. We therefore started searching for beekeepers  
 
that would agree to discuss with us.
 
that would agree to discuss with us.
 
  </p>
 
  </p>
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   <div class="group center">
 
   <div class="group center">
−
  <p align="justify" style="font-size:15px;">In conclusion of this meeting we decided that  
+
  <p align="justify" style="font-size:15px;">The conclusion of this meeting was that  
 
  the bacteria should not produce both attractive and toxic molecules at the same time. These  
 
  the bacteria should not produce both attractive and toxic molecules at the same time. These  
−
  productions should be cyclic to have the least effect on bee’s life cycle since formate is also toxic at a lesser extent for bees.
+
  productions should be cyclic to have the least effect on bees life cycle since formate is also toxic at a lesser extent for bees.
 
  </p>
 
  </p>
 
  </div>
 
  </div>
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  <div class="group center">
 
  <div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
  During the whole week we searched for means to produce our molecules cyclically. We found a way to produce our molecules
+
  During the whole week we searched for ways to produce our molecules cyclically. We chose to implement
−
  on a circadian rhythm which allows us to produce toxic molecules during the night when bees are a lot less active.<br>
+
  a circadian rhythm cycle which would allow us to produce toxic molecules during the night when bees are a lot less active.<br>
−
Since we agreed on the regulation constructions we started to look for means to build our construction.<br>
+
  We also convinced Louise Gody to create our logo because she is capable of drawing with talent using a graphical tablet.<br>
−
  We also started to think about our logo and decided that Louise Gody will create our logo since she is capable of drawing using a graphical tablet.<br>
+
We also started to organise our search for financial help. To do so we designed a booklet explaining our project and what iGEM is.<br>
−
We also started to organize our search for financial help. To do so we designed a booklet explaining our project and what iGEM is.<br>
+
 
  </p>
 
  </p>
 
  </div>
 
  </div>
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   <div class="group center">
 
   <div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
We started searching for each individual genes by first looking on the iGEM registry.<br>
+
We first started searching for genes that we would like to have in our constructions on the iGEM registry.<br>
−
Knowing that the design of our construct was agreed upon we also started thinking about what our device would look like. Since our bacteria needed to perceive red light we decided to place our device at the entry of the hive where bees need to enter the beehive.
+
After we successfully designed our construct we started thinking about what our device would look like. Since our bacteria needed to perceive red light we decided to place our device at the entry of the hive but outside.
 
  </p>
 
  </p>
 
  </div>
 
  </div>
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   <div class="group center">
 
   <div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
  We realised that our work would be too heavy seeing the number of genes needed to obtain and the cloning work required.. We therefore decided to synthesise the sequence needed to realise our construction.
+
  We realised that our the cloning part would be quite long because of the number of genes needed to obtain... We therefore decided to order synthetic sequences for our synthetic pathway constructions.
 
  </p>
 
  </p>
 
  </div>
 
  </div>
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   <div class="group center">
 
   <div class="group center">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
Laetitia Chaumont, Benoit Pons and Mathilde Beraud went to Vacbio EA 4357 a laboratory studying varroa. Dr Vetillard allowed us to test the actual effects different concentrations of butyrate on Varroa. The results were inconclusive. However two parameters should be improved because butyrate is a very volatile molecule and we conducted experiments in a petri dish, that is why we decided to set up a more optimised protocol to assess the attraction of varroa by butyrate.
+
Laetitia Chaumont, Benoit Pons and Mathilde Beraud went to a laboratory studying varroa(Vacbio EA 4357). Dr. Vetillard allowed us to test the actual effects of different concentrations of butyrate on Varroa. The results were inconclusive. However two parameters had to be improved because butyrate is a very volatile molecule and we conducted experiments in a petri dish thus diffusion complicated the interpretation. Therefore, we decided to set up a more optimised protocol to assess the attraction of varroa by butyrate.
 
  </p>
 
  </p>
 
  </div>
 
  </div>
 +
 +
<div class="subsubtitle">
 +
<h3>06/18</h3>
 +
</div>
 +
 +
<div class="group">
 +
<p align="justify" style="font-size:15px;">
 +
Louise Gody drew our amazing logo!! <br><br> </p> </div>
 +
<center><img src=" https://static.igem.org/mediawiki/2015/f/f8/TLSE_ApiColi_Logo.png" style="width:33%;" /></center>
 +
 
   
 
   
 
  <div class="subsubtitle">
 
  <div class="subsubtitle">
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  <br>
 
  <br>
 
   <ul align="justify" style="font-size:15px;">
 
   <ul align="justify" style="font-size:15px;">
−
<li> Modeling started. We attempted to determine which metabolic pathways our molecules are taking part. This gave us the opportunity to have an overview of their roles and effects. </li>
+
<li> Modelling started. We attempted to determine which metabolic pathways our molecules are taking part. This gave us the opportunity to have an overview of their roles and effects. </li>
 
<li>Everybody trained to master the different protocols of synthetic biology. Our constructs were finally ordered via Biobasic which offered us a discount price.</li>
 
<li>Everybody trained to master the different protocols of synthetic biology. Our constructs were finally ordered via Biobasic which offered us a discount price.</li>
  
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  <div class="group">
 
  <div class="group">
 
  <p align="justify" style="font-size:15px;">
 
  <p align="justify" style="font-size:15px;">
−
  We designed a new protocol to assess whether butyrate is an attractive molecule for varroas. Thanks to M. Patrick Chekroun who made a glass T-tube, we were able to define the attraction power of butyrate. <a target="_blank" href="https://2015.igem.org/Team:Toulouse/Experiments">See "Protocols" part</a>
+
  We designed a new protocol to assess whether butyrate is an attractive molecule for varroas. Thanks to M. Patrick Chekroun who made a T-tube in glass, we were able to define the attraction power of butyrate. <a target="_blank" href="https://2015.igem.org/Team:Toulouse/Experiments">See "Protocols" part</a>
 
</p>
 
</p>
 
  </div>
 
  </div>
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  <div class="group">
 
  <div class="group">
 
  <p align="left" style="font-size:15px;">
 
  <p align="left" style="font-size:15px;">
−
  Our project was in a popular newspaper “Metronews”.
+
  Our project was described in a popular newspaper “Metronews”.
 
  </p>
 
  </p>
 
  </div>
 
  </div>
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  <div class="group">
 
  <div class="group">
 
  <p align="left" style="font-size:15px;">
 
  <p align="left" style="font-size:15px;">
−
Our project made the cover of a popular local newspaper “La Dépèche”.
+
Our project made the cover of a popular local newspaper “La Dépêche”.
 
  </p>
 
  </p>
 
  </div>  
 
  </div>  
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  <div class="group center">
 
  <div class="group center">
−
   <img src=" https://static.igem.org/mediawiki/2015/3/32/TLSE_Notebook_3.png" style="width:30%;height:auto;" />
+
   <img src=" https://static.igem.org/mediawiki/2015/3/32/TLSE_Notebook_3.png" style="width:30%;" />
 
   </div>
 
   </div>
 
  </center>
 
  </center>
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   <div class="group">
 
   <div class="group">
 
  <p align="left" style="font-size:15px;">
 
  <p align="left" style="font-size:15px;">
−
  First competent cells success by Benoît Pons!!   
+
  First competent cells obtained with success by Benoît Pons!!   
 
   </p>
 
   </p>
 
  </div>  
 
  </div>  
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  <br>
 
  <br>
 
<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
−
<li> We Skyped with the iGEM Ankara team which is also working on Varroa destructor. We exchanged on our different strategies, and decided to start a collaboration with us in order to combine our efforts to fight against varroa and to gather some data about the mite. See <a target="_blank" href="https://2015.igem.org/Team:Toulouse/Collaborations#ankara">  "Collaborations" </a>  part for more information.  </li>
+
<li> We Skyped with the iGEM Ankara team which is also working on <i>Varroa destructor</i>. We exchanged on our different strategies, and decided to start a collaboration with them. Indeed, we wanted to combine our efforts to fight against varroa and to gather some data about the mite. See <a target="_blank" href="https://2015.igem.org/Team:Toulouse/Collaborations#ankara">  "Collaborations" </a>  part for more information.  </li>
 
<li>We stored two bacterial strains BW25113 and MG1665 in a glycerol solution. </li>
 
<li>We stored two bacterial strains BW25113 and MG1665 in a glycerol solution. </li>
 
</ul>
 
</ul>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
Following the newspaper articles published we discovered a forum of beekeepers discussing about our project on the web. A lot of them were perplexed by our project. We decided to engage the conversation with them and answer all of their questions.
+
Following the newspaper articles published we discovered a forum of beekeepers discussing about our project on the web. A lot of them were puzzled by our project. We decided to engage the conversation with them and answer all of their questions. They were pleasantly surprised that we took the time to answer all their questions!
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
We prepared precursor molecules for a M9 medium, and also did a cytotoxicité test of formic acid thanks to a plate reader.  
+
We prepared some M9 medium, and also did a cytotoxicity test of formic acid thanks to a plate reader.  
 
</p>
 
</p>
 
</div>
 
</div>
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     <div class="group">
 
     <div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
We realised two tests: one is a test culture on BW25113 strain on aerobic and micro aerobic conditions. The other permits to determine the cytotoxicity of formic acid.
+
We realised two tests: one is a test culture on BW25113 strain under aerobic and micro aerobic conditions. The other permits to determine the cytotoxicity of formic acid.
 
</p>
 
</p>
 
</div>
 
</div>
 
 
 
<center>
 
<center>
−
  <div class="group center">
+
   
−
   <img src=" https://static.igem.org/mediawiki/2015/0/0d/TLSE_Notebook_6.jpg" style="width:40%;height:auto;" />
+
   <img src=" https://static.igem.org/mediawiki/2015/0/0d/TLSE_Notebook_6.jpg" style="width:40%;" />
−
  </div>
+
 
 
  </center>
 
  </center>
 
   
 
   
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<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
 
<li> Laetitia Chaumont obtained a meeting with Vincent Grégoire Delory, the ethician of Toulouse White Biotechnology (TWB).  </li>
 
<li> Laetitia Chaumont obtained a meeting with Vincent Grégoire Delory, the ethician of Toulouse White Biotechnology (TWB).  </li>
−
<li> We did a bacterial culture monitoring thanks to an Optical Density (OD) reading and culture plating. </li>
+
<li> We performed a bacterial culture growth monitoring. </li>
 
<li>We did again a cytotoxicity test of formic acid. </li>
 
<li>We did again a cytotoxicity test of formic acid. </li>
 
  </ul>
 
  </ul>
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<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
−
<li> We did a bacterial culture monitoring thanks to an Optical Density (OD) reading and culture plating.  </li>
+
<li> Again a bacterial culture growth monitoring.  </li>
−
<li> We did a cytotoxicity test of butyric acid. </li>
+
<li> Cytotoxicity test of butyric acid done. </li>
 
</ul>
 
</ul>
 
 
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<br>
 
<br>
 
   <ul align="justify" style="font-size:15px;">
 
   <ul align="justify" style="font-size:15px;">
−
<li> We did a bacterial culture monitoring thanks to an Optical Density (OD) reading and culture plating.  </li>
+
<li> Bacterial culture growth monitoring (again!!).  </li>
−
<li> We did a cytotoxicity test of butyric acid. </li>
+
<li> We also realised a cytotoxicity test of butyric acid (again!!). </li>
 
   
 
   
 
</ul>
 
</ul>
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   <ul align="justify" style="font-size:15px;">
 
   <ul align="justify" style="font-size:15px;">
 
<li> We did cloning trainings to be as efficient as possible, Benoît Pons and Marine Pons succeeded the first cloning at the first attempt !!    </li>
 
<li> We did cloning trainings to be as efficient as possible, Benoît Pons and Marine Pons succeeded the first cloning at the first attempt !!    </li>
−
<li> We did a bacterial culture monitoring thanks to an Optical Density (OD) reading and culture plating. </li>
+
<li> Bacterial culture growth monitoring (again!!). </li>
−
<li>We did a cytotoxicity test of butyric acid. </li>
+
<li>We also realised a cytotoxicity test of butyric acid (again!!).</li>
 
   
 
   
 
</ul>
 
</ul>
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<br>
 
<br>
 
   <ul align="justify" style="font-size:15px;">
 
   <ul align="justify" style="font-size:15px;">
−
<li> We did a bacterial culture monitoring thanks to an Optical Density (OD) reading and culture plating. </li>
+
<li>Bacterial culture growth monitoring (again!!). </li>
−
<li>We did a cytotoxicity test of butyric acid. </li>
+
<li>We also realised a cytotoxicity test of butyric acid (again!!).</li>
 
   
 
   
 
</ul>
 
</ul>
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<br>
 
<br>
 
   <ul align="justify" style="font-size:15px;">
 
   <ul align="justify" style="font-size:15px;">
−
<li> All the team was waiting for the synthetic genes and training since 26th june with some basic technics such as miniprep, gel extraction, transformation, cloning, cells culture. We started to be pessimists about the next phase of the project, but we kept a good mood in the team.</li>
+
<li> All the team is now waiting for the synthetic genes and training since the 26th of June. Training with some basic technics such as miniprep, gel extraction, transformation, cloning, cells culture. We are eager to start, but are a bit pessimistic about the next phase of the project. Nonetheless, good mood of the team!!</li>
−
<li>We did a bacterial culture monitoring thanks to an Optical Density (OD) reading and culture plating. </li>
+
<li>Bacterial culture growth monitoring (again!!).   </li>
−
<li>We did a cytotoxicity test of butyric acid. </li>
+
<li>We also realised a cytotoxicity test of butyric acid (again!!).</li>
 
   
 
   
 
</ul>
 
</ul>
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<center>
 
<center>
−
<div class="group center">
+
 
−
   <img src="https://static.igem.org/mediawiki/2015/0/08/TLSE_Protocols_Culture_4.png" style="width:25%;height:auto;" />
+
   <img src="https://static.igem.org/mediawiki/2015/0/08/TLSE_Protocols_Culture_4.png" style="width:25%;" />
−
  </div>
+
 
 
  </center>
 
  </center>
  
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
We received from from Dr Nico Claasens lab (Wageningen University) the strains containing PCB en Cph8 vectors responsibles for the circadian regulation. We could now begin the genetic construction, but we faced to a compatibility vectors problem. Indeed, receptors genes were cloned into vectors incompatible with our final construction plan, so we had to look for a solution.
+
We received from Dr Nico Claasens lab (Wageningen University) the strains containing PCB en Cph8 vectors responsible for the circadian regulation. We could now begin the genetic construction, but we faced a problem with the vectors: receptors genes were cloned into vectors incompatible with our final construction plan, so we had to look for an alternative solution.
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
After the ordering of our synthetic genes, we were waiting for them. <br> We decided to work in pairs per module to be as effective as possible:
 
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
We received our synthetic genes after more than one month of waiting. There is only one month left and we had to work harder to reach project goals.  Now the real fight could begin, hoping to achieve that we planed.
+
We received our synthetic genes after more than one month of waiting for them. There is only one month left and we have to work harder to reach project goals.  Now the real fight begins, hoping to achieve that we planed.
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
The synthetic genes, which were cloned into pUC57 vectors, were transformed into E. coli strain in order to realize miniprep and then cloned during the weekend. We were excited to experiment something new, and see if all our efforts spent on gene design paid off.
+
The synthetic genes, which were cloned into pUC57 vectors, were amplified and then cloned during the weekend. We were excited to experiment something new, and see if all our efforts spent on gene design paid off.
 
</p>
 
</p>
 
</div>
 
</div>
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<br>
 
<br>
 
<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
−
<li>Miniprep of bacteria containing synthetical genes, followed by a digestion by NotI restriction enzyme of the plasmid containing the formate genes (pflA and pflB). Then a ligation of puc57 wad done in ordr to obtain the empty vector. Finally, a transformation was realized and the results were good. Indeed, there was no colony in the negative control  plate, many colonies on the positive control and one clone for our ligation. </li>
+
<li>Miniprep of the clones containing the synthetic genes.</li>
−
<li>First miniprep, digestion, gel extraction, cloning and transformation were performed for all the different modules and we are eager to see clones on LB plates. </li>
+
<li>First miniprep, digestion, gel extraction, cloning and transformation were performed for all the different modules. </li>
 
</ul>
 
</ul>
  
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<br>
 
<br>
 
<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
−
<li>We had some cloning which worked and others which did not. To check that it really worked for some, we decided to do a miniprep.Indeed, if the insert has been cloned into the vector, the plasmid will be heavier. So we had to do cells culture with some clones. </li>
+
<li>Some cloning worked and others did not. Verification of the clones. Start again for the ones that did not work...</li>
−
<li>For the clonings which did not work, we had to do them again. </li>
+
−
<li>We checked the only clone obtained the 23/08 and we concluded that it corresponds to puc57. </li>
+
−
<li>Recircularized puc57 vector became the vector control for culture tests.  </li>
+
−
</ul>
+
  
  
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<br>
 
<br>
 
<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
−
<li>The first clonings worked for the final construction: a first step has been completed. Unfortunately we lost a teammate in this battle, because Anthony had to leave the project in order to start his courses in Sweden.  </li>
+
<li>The first cloning worked for the final construction: a first step has been completed (the formate genes were cloned into the PL-CI pUC57 plasmid). Unfortunately we lost a team mate in this battle, because Anthony had to leave the project in order to start his courses in Sweden.  </li>
 
<li>Tests of formate production were made in Falcon tubes in order to mimic micro aerobic conditions. </li>
 
<li>Tests of formate production were made in Falcon tubes in order to mimic micro aerobic conditions. </li>
 
<li>Culture tests were realized for different concentrations of the Biosilta medium.    </li>
 
<li>Culture tests were realized for different concentrations of the Biosilta medium.    </li>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
The formic culture was followed.  
+
The production of formic acid in the culture of the producing strain was followed.  
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
Preparation of NMR tubes.  
+
Preparation of NMR samples and analysis.  
 
</p>
 
</p>
 
</div>
 
</div>
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<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
 
<li><strong>Success</strong> of the PcyA-tesB cloning  </li>
 
<li><strong>Success</strong> of the PcyA-tesB cloning  </li>
−
<li>French institution called DRAAF announced us that they support us with <strong>€ 3000!! </strong>  </li>
+
<li>French institution called DRAAF announced us that they support us with <strong>€ 3000!! </strong>  </li>
 
</ul>
 
</ul>
  
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
  Anthony met Felix from iGEM Stockholm Team, and discussed about the iGEM experience, all the work done at this moment, his disappointments, … He understood that all iGEM teams were in the same situation, because of time lack.
+
  Anthony met Felix from iGEM Stockholm Team, and discussed about the iGEM experience, all the work done at this moment, his disappointments, … He understood that all iGEM teams were in the same situation: URGENCY.
 
</p>
 
</p>
 
</div>
 
</div>
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
  We succeeded <strong>the first In Fusion cloning</strong> and the crt-ho1 cloning.  
+
  We succeeded <strong>the first In Fusion cloning step</strong> and the cph8 cloning.  
 
</p>
 
</p>
 
</div>
 
</div>
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<br>
 
<br>
 
<ul align="justify" style="font-size:15px;">
 
<ul align="justify" style="font-size:15px;">
−
<li>Even if the experiments did not work as expected, all the team smiles again when we received our polo shirts and berets that we will wear in Boston: we were ready to fight !  </li>
+
<li>Even if the experiments did not work as expected, all the team smiles again when we received our polo shirts and berets that we will wear in Boston: we are ready to fight !  </li>
−
<li>Another good new was the success of circadian clonings made by Benoît Pons </li>
+
<li>Another good news was the success of the first steps of the circadian cloning made by Benoît Pons</li>
 
</ul>
 
</ul>
  
 
<center>
 
<center>
−
<div class="group center">
+
 
−
   <img src="https://static.igem.org/mediawiki/2015/5/56/TLSE_Notebook_5.jpg" style="width:60%;height:auto;" />
+
   <img src="https://static.igem.org/mediawiki/2015/5/56/TLSE_Notebook_5.jpg" style="width:60%;" />
−
  </div>
+
 
  </center>
 
  </center>
  
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<div class="group">
 
<div class="group">
 
<p align="justify" style="font-size:15px;">
 
<p align="justify" style="font-size:15px;">
−
  This day, we succeeded the cloning of ccr-butyrate (done the 29/08) in pSB1C3.  
+
  This day, we succeeded the cloning of ccr-butyrate (done the 08/29), PL-LacI-ccr-Butyrate genes (done the 08/28) and P<sub>OmpC</sub>-lacI box-CI-Formate genes (done the 08/25) into pSB1C3.  
 +
 
 
</p>
 
</p>
 
</div>
 
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+
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<h3>09/19</h3>
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 +
<div class="group">
 +
<p align="justify" style="font-size:15px;">
 +
Working past midnight in the lab ? Strong headaches and itching eyes but still on the computer ? Fighting with HTML tags and continuously looking at the clock ? No doubt, this is the final effort before the dreadful wikifreeze !!!
 +
</p>
 +
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Latest revision as of 22:15, 18 September 2015

iGEM Toulouse 2015

Notebook


Calendar - 2015

January

During this period, we were informed by iGEM toulouse instructors of what this competition is, and what was expected of us.

From February to May

02/02

First team meeting, our instructors informed us on the competition more specifically and how the team is organized. Our first mission was to prepare a presentation for the next week on the following themes:

  • Previous subjects. What are the themes explored by iGEM? Give examples of good projects and explain why they won?
  • Biobricks? What are they? What are their advantages? What are the materials sent by iGEM? How does the registry work?
  • Competition and deadlines. How does the competition works and what are the important deadlines?

Those presentations gave us the opportunity to understand more about the iGEM competition and to get to know each other (since we don’t all come from the same institutions).

Our first team meeting

We also created our email address and a google drive to organise our files online.

02/09


Every group of two or three students presented one of the subjects seen above. Every presentation was followed by a Q&A. Thanks to it, everyone understood clearly the topic.

02/16

The next step was to present every week, two new project ideas. The presentations were divided in the following parts.

  • Context
  • Goal
  • Originality
  • Strategy
  • Difficulty
  • Modelling
  • Device
  • Benefit
  • Ethics
  • Planning
  • Why is this project going to win?

Until May

The following weeks were spent brainstorming. Everyone presented at least two project ideas. Here are few examples of projects reviewed:

  • Creation of a quick detection kit of hepatitis
  • Degradation of Lignin
  • Use of ice nucleation proteins for water desalination
  • Creation of a small domestic pollution detector
  • ...

May

05/04

Our youngest instructor and team mother Mathilde Béraud went with Benoît Pons to the FSIE committee, and thanks to their intervention, we obtained € 4000 !

05/05

Two projects were still in course to be chosen, the use of ice nucleating proteins to purify water and the creation of a bio trap against the parasitic mite Varroa destructor. This last one was chosen to be our project on the grounds that it was the most feasible. Based on bibliography, the initial idea was to build an engineered bacteria which will have to produce butyrate to attract the parasite. The following weeks were the subject of extensive researches on the topic.

Moreover, it was time to choose attributions to make the project management easier, below in the table, the main assignments.

Trouche Blandine Finance
Tanchon Melany Modelling
David Melissa Events
Pons Benoit Press
Etcheberry Thomas Logistics
Le Scornet Alexandre Registry, Biobricks
Pons Marine Experiments
David Anthony Power Point Presentation
Gody Louise Wiki
Chaumont Laetitia Ethics

05/13

Two members of our team participated in the Toulouse Exposciences. They presented a poster on synthetic biology and the iGEM competition. They also organized banana DNA extraction experiences for children. This successful event was a good opportunity for us to start learning how to design a poster and to communicate about synthetic biology.

05/15

We held our first meeting without instructors to discuss the design of the constructions needed to create the engineered bacteria. Our reflection led us to the conclusion that our construction should respect the bee life cycle. We therefore started searching for beekeepers that would agree to discuss with us.

05/18

We met with Dr Boucher Christian HDR, DR1 INRA retired since 2012 and amateur beekeeper. The fact that Dr Boucher was both a researcher and a beekeeper made this meeting really beneficial for us. We learned every important stages in a bee life and how the varroa affected it. We also learned more about the beekeeping activity and the treatments used to fight against the parasite. Furthermore we have been able to discuss our early genetic design and how to improve it.

The conclusion of this meeting was that the bacteria should not produce both attractive and toxic molecules at the same time. These productions should be cyclic to have the least effect on bees life cycle since formate is also toxic at a lesser extent for bees.

05/25

During the whole week we searched for ways to produce our molecules cyclically. We chose to implement a circadian rhythm cycle which would allow us to produce toxic molecules during the night when bees are a lot less active.
We also convinced Louise Gody to create our logo because she is capable of drawing with talent using a graphical tablet.
We also started to organise our search for financial help. To do so we designed a booklet explaining our project and what iGEM is.

June

06/01

We first started searching for genes that we would like to have in our constructions on the iGEM registry.
After we successfully designed our construct we started thinking about what our device would look like. Since our bacteria needed to perceive red light we decided to place our device at the entry of the hive but outside.

06/05

We realised that our the cloning part would be quite long because of the number of genes needed to obtain... We therefore decided to order synthetic sequences for our synthetic pathway constructions.

06/06

Laetitia Chaumont, Benoit Pons and Mathilde Beraud went to a laboratory studying varroa(Vacbio EA 4357). Dr. Vetillard allowed us to test the actual effects of different concentrations of butyrate on Varroa. The results were inconclusive. However two parameters had to be improved because butyrate is a very volatile molecule and we conducted experiments in a petri dish thus diffusion complicated the interpretation. Therefore, we decided to set up a more optimised protocol to assess the attraction of varroa by butyrate.

06/18

Louise Gody drew our amazing logo!!

06/22

We finished the design of our constructions and send it to our instructors for checking. In parallel we started training ourselves on basic synthetic biology protocols such as making competent cells, biobrick transformation…

06/29


  • Modelling started. We attempted to determine which metabolic pathways our molecules are taking part. This gave us the opportunity to have an overview of their roles and effects.
  • Everybody trained to master the different protocols of synthetic biology. Our constructs were finally ordered via Biobasic which offered us a discount price.


July

07/01

We designed a new protocol to assess whether butyrate is an attractive molecule for varroas. Thanks to M. Patrick Chekroun who made a T-tube in glass, we were able to define the attraction power of butyrate. See "Protocols" part

07/02

Our project was described in a popular newspaper “Metronews”.

07/04

Our project made the cover of a popular local newspaper “La Dépêche”.

07/06


  • Laetitia Chaumont initiated cytotoxicity test. The purpose of those tests are to assess whether our bacteria could withstand various butyrate/formate concentration
  • In parallel Melany Tanchon determined the production of butyrate/formate. Those results are directly linked to the undergoing cytotoxicity tests.
  • We also initiated the creation of the plans that would be used to 3D print our device. In order to achieve this, Valentin Girin used the software Catia for the modeling of our device.
  • Our crowdfunding campaign was launched via the Ulule platform.

07/08

First competent cells obtained with success by Benoît Pons!!

07/09


  • We Skyped with the iGEM Ankara team which is also working on Varroa destructor. We exchanged on our different strategies, and decided to start a collaboration with them. Indeed, we wanted to combine our efforts to fight against varroa and to gather some data about the mite. See "Collaborations" part for more information.
  • We stored two bacterial strains BW25113 and MG1665 in a glycerol solution.

07/13

Following the newspaper articles published we discovered a forum of beekeepers discussing about our project on the web. A lot of them were puzzled by our project. We decided to engage the conversation with them and answer all of their questions. They were pleasantly surprised that we took the time to answer all their questions!

07/14

We prepared some M9 medium, and also did a cytotoxicity test of formic acid thanks to a plate reader.

07/16

We realised two tests: one is a test culture on BW25113 strain under aerobic and micro aerobic conditions. The other permits to determine the cytotoxicity of formic acid.

48 wells plate, different formic acid concentrations

07/17


  • Laetitia Chaumont obtained a meeting with Vincent Grégoire Delory, the ethician of Toulouse White Biotechnology (TWB).
  • We performed a bacterial culture growth monitoring.
  • We did again a cytotoxicity test of formic acid.

07/18

  • Again a bacterial culture growth monitoring.
  • Cytotoxicity test of butyric acid done.

07/20

Melissa David, Melany Tanchon and Louise Gody went to Brussels to attend the international Exposciences for the whole week. They presented our project and synthetic biology to people coming from all around the world.

07/21 & 07/22


  • Bacterial culture growth monitoring (again!!).
  • We also realised a cytotoxicity test of butyric acid (again!!).

07/23


  • We did cloning trainings to be as efficient as possible, Benoît Pons and Marine Pons succeeded the first cloning at the first attempt !!
  • Bacterial culture growth monitoring (again!!).
  • We also realised a cytotoxicity test of butyric acid (again!!).

07/24


  • Bacterial culture growth monitoring (again!!).
  • We also realised a cytotoxicity test of butyric acid (again!!).

07/27


  • All the team is now waiting for the synthetic genes and training since the 26th of June. Training with some basic technics such as miniprep, gel extraction, transformation, cloning, cells culture. We are eager to start, but are a bit pessimistic about the next phase of the project. Nonetheless, good mood of the team!!
  • Bacterial culture growth monitoring (again!!).
  • We also realised a cytotoxicity test of butyric acid (again!!).

07/29

Preparation of NMR tubes for the analysis of supernatants.

07/30

We did a cytotoxicity test of butyric acid and we were trained to be able to use the NMR machine.

NMR training

07/31

An enzymatic test was done on the Biosilta kit with a plate reader.

August

08/03

Culture tests started !

08/04


  • We did a very good glucose dosage.
  • An enzymatic test was done on the Biosilta kit with an OD reading, until the stabilization of bacterial growth.

08/05

We received from Dr Nico Claasens lab (Wageningen University) the strains containing PCB en Cph8 vectors responsible for the circadian regulation. We could now begin the genetic construction, but we faced a problem with the vectors: receptors genes were cloned into vectors incompatible with our final construction plan, so we had to look for an alternative solution.

08/15

  • Thomas and Mélany worked together on the attraction module: production of butyrate
  • Laetitia worked on the eradication module: production of formate
  • Benoît developed the regulation module
  • Marine and Anthony worked on the final construction
  • Alexandre and Mélissa did the different biobricks

08/17

Preparation of NMR samples.

08/19

Enzymatic tests on glucose oxydase.

08/20

We received our synthetic genes after more than one month of waiting for them. There is only one month left and we have to work harder to reach project goals. Now the real fight begins, hoping to achieve that we planed.

08/22

The synthetic genes, which were cloned into pUC57 vectors, were amplified and then cloned during the weekend. We were excited to experiment something new, and see if all our efforts spent on gene design paid off.

08/23


  • Miniprep of the clones containing the synthetic genes.
  • First miniprep, digestion, gel extraction, cloning and transformation were performed for all the different modules.

08/24


  • Some cloning worked and others did not. Verification of the clones. Start again for the ones that did not work...

    • 08/25


    • The first cloning worked for the final construction: a first step has been completed (the formate genes were cloned into the PL-CI pUC57 plasmid). Unfortunately we lost a team mate in this battle, because Anthony had to leave the project in order to start his courses in Sweden.
    • Tests of formate production were made in Falcon tubes in order to mimic micro aerobic conditions.
    • Culture tests were realized for different concentrations of the Biosilta medium.

    08/26 & 08/27

    The production of formic acid in the culture of the producing strain was followed.

    08/28

    The formate genes (pflA and pflB), the butyrate genes without ccr and the POmpC-LacI box-CI were cloned into pSB1C3. The butyrate genes were cloned into the PL-LacI-ccr pUC57 Plasmid.

    08/29


    • Blandine took part to iGEM meetup France organized by iGEM Bordeaux Team, where she met different teams, discussing with them, and presenting our project.
    • This same day, we succeeded the cloning of ccr-butyrate together.

    08/30

    Preparation of NMR samples and analysis.

    September

    09/01


    • Success of the PcyA-tesB cloning
    • French institution called DRAAF announced us that they support us with € 3000!!

    09/04

    Anthony met Felix from iGEM Stockholm Team, and discussed about the iGEM experience, all the work done at this moment, his disappointments, … He understood that all iGEM teams were in the same situation: URGENCY.

    09/07

    We succeeded the first In Fusion cloning step and the cph8 cloning.

    09/10


    • Even if the experiments did not work as expected, all the team smiles again when we received our polo shirts and berets that we will wear in Boston: we are ready to fight !
    • Another good news was the success of the first steps of the circadian cloning made by Benoît Pons

    09/12

    Blandine, who created our Ulule account (Ulule is a crowdfunding platform) informed us that it was a real success. Indeed, during the crowdfunding period, we obtained € 1,780, with the support of 59 contributors.

    09/15

    This day, we succeeded the cloning of ccr-butyrate (done the 08/29), PL-LacI-ccr-Butyrate genes (done the 08/28) and POmpC-lacI box-CI-Formate genes (done the 08/25) into pSB1C3.

    09/16

    All Biobricks were prepared and sent to be sequenced

    09/17

    One day before THE deadline we sent our Biobrick to iGEM !!

    09/19

    Working past midnight in the lab ? Strong headaches and itching eyes but still on the computer ? Fighting with HTML tags and continuously looking at the clock ? No doubt, this is the final effort before the dreadful wikifreeze !!!