Difference between revisions of "Team:HUST-China/Model"

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     <title>Attributions</title>
 
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                    <li><a href="https://2017.igem.org/Team:HUST-China">HOME</a></li>
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                    <li><a href="https://2017.igem.org/Team:HUST-China">HOME</a></li>
 
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                             WETLAB <b class="caret"></b>
 
                             WETLAB <b class="caret"></b>
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                             TEAM <b class="caret"></b>
 
                             TEAM <b class="caret"></b>
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                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
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     <div class="tab1" style="width:100%;margin-top: 65px;">
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     <div class="tab1" id="section0" style="width:100%;margin-top: 65px;">
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             <h1 class="tab-h1" ><strong>「Experiment」</strong></h1>
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             <h1 class="tab-h1" ><strong>「Attributions」</strong></h1>
 
     </div>
 
     </div>
  
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                 <div class="collapse navbar-collapse" id="navbar-collapse">
 
                 <div class="collapse navbar-collapse" id="navbar-collapse">
 
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                 <ul class="nav nav-pills nav-stacked" style="font-size:12px; text-align:center; background:#ccc;">
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                     <li class="active"><a href="#section1">Sensing part</a></li>
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                     <li class="active"><a href="#section1">Overview</a></li>
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                     <li><a href="#section2">Recycling part</a></li>
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                     <li><a href="#section2">Assumptions</a></li>
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                     <li><a href="#section3">The whole circuit </a></li>
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                     <li><a href="#section3">Constant & Parameter</a></li>
 +
                    <li><a href="#section4">Basical Diffusion</a></li>
 +
                    <li><a href="#section5">Recycling</a></li>
 +
                    <li><a href="#section6">Sensing and Capture</a></li>
 +
 
 
                 </ul>
 
                 </ul>
 
                 </div>   
 
                 </div>   
 
                 </div>       
 
                 </div>       
 
             </nav>
 
             </nav>
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             <div class="col-sm-8 text">
 
                 <div id="section1" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">     
 
                 <div id="section1" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">     
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                     <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px;margin-top:0;">Sensing part</h3>
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                     <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px;margin-top:0;">Overview </h3>
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                     <p>
+
                     <p>We build an ecosystem level model, in order to see how the Reebot diffuse, sense and capture the lanthanide ion, and be recycled on Si-board. Then we can determine how should we release Reebot, how to recycle the Reebot and how much Reebot do we need to produce the best result. The ecosystem could derive the best strategy to make full use of Reebot.
−
                        The sensing part is to achieve the awareness of the engineering bacteria on the rare earth ions in the environment.
+
 
                     </p>
 
                     </p>
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                    <h4><strong>①Amplify</strong> </h4>
+
                </div>
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                    <p>
+
−
                        pmrA-B-Fe、pmrC/GFP  gene through the pcr protocol
+
−
                    </p>
+
−
                    <h4><strong>②Plasmid Construction</strong> </h4>
+
−
                    <p>
+
−
                      Coding sequences pmrA/pmrB and pmrC-GFP were cloned into the expression vector pBAD30 by restriction enzyme digestion and DNA ligation.
+
−
                    </p>
+
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                    <div class="container experiment_circuit col-md-6 col-xs-6 col-md-offset-3 col-xs-offset-3">
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                        <img src="https://static.igem.org/mediawiki/2017/d/d5/2017_HUST_China_Experiment_image1.png" alt="experiment_circuit" class="img-responsive">
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                    </div>
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                    <div class="row col-xs-12">
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                        <p>
+
−
                            Then we transformed these vectors into Ecoli BL21 cells.
+
−
                        </p>
+
−
                        <h4><strong>③Detection</strong> </h4>
+
−
                        <p>
+
−
                          Arabinose was used to induce expression of pmrABC system.
+
−
                        </p>
+
−
                        <p>
+
−
                          We use the original pmrA-pmrB / Fe-pmrC-GFP circuit to test whether our
+
−
                          pmrABC system works in our plasmid, then the expression and efficacy
+
−
                          of LBT1-12 were tested .
+
−
                        </p>
+
−
                    </div>
+
  
 +
                <div id="section2" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">
 +
                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Assumptions</h3>
 +
                    <p>The variability of temperature has no effect on our Reebot or it could be ignored.</p>
 +
                    <p>Liquid in the container is stationary.</p>
 +
                    <p>There is nothing else in the container which can influence Reebot.</p> 
 +
                </div>
  
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                     <div class="table-responsive" style="padding: 10px 100px; text-align: center; font-size:14px;">
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                <div id="section3" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">       
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                         <table class="table">
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                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Constant & Parameter</h3>
 +
                    <p>The description of constant, their values and the references involved in this model are listed in this table.</p>
 +
 
 +
                     <div class="table-responsive " style="padding: 10px 100px; text-align: center; font-size:14px;">
 +
                         <table class="table table-hover">
 
                           <thead>
 
                           <thead>
 
                               <tr>
 
                               <tr>
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                                 <th style="text-align: center;">gene circuit</th>
+
                                 <th style="text-align: center;">Constant </th>
−
                                 <th style="text-align: center;">vector</th>
+
                                 <th style="text-align: center;">Description</th>
−
                                <th style="text-align: center;">description</th>
+
 
                               </tr>
 
                               </tr>
 
                           </thead>
 
                           </thead>
 +
 +
 
                           <tbody>
 
                           <tbody>
 
                                   <tr>
 
                                   <tr>
−
                                     <td>pmrA-pmrB/Fe-pmrC-GFP</td>
+
                                     <td>f<sub>1</sub></td>
−
                                    <td>pBAD30</td>
+
                                     <td>concentration of Initially Bacteria</td>
−
                                     <td>Test whether our pmrABC system works in our plasmid</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT1-pmrC-GFP </td>
+
                                    <td>f<sub>2</sub></td>
−
                                    <td>pBAD30</td>
+
                                    <td>concentration of Lanthanide ion</td>
−
                                    <td> Detecting effectiveness of protein LBT1</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT2-pmrC-GFP</td>
+
                                    <td>f<sub>3</sub></td>
−
                                    <td>pBAD30 </td>
+
                                    <td>concentration of Sensed Bacteria</td>
−
                                    <td> Detecting effectiveness of protein LBT2</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT3-pmrC-GFP</td>
+
                                    <td>f<sub>4</sub></td>
−
                                    <td>pBAD30 </td>
+
                                    <td>concentration of Captured Bacteria</td>
−
                                    <td>Detecting effectiveness of protein LBT3</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT4-pmrC-GFP</td>
+
                                    <td>f<sub>5</sub></td>
−
                                    <td> pBAD30 </td>
+
                                    <td>concentration of recycled sensed bacteria </td>
−
                                    <td>Detecting effectiveness of protein LBT4</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT5-pmrC-GFP</td>
+
                                    <td>f<sub>6</sub></td>
−
                                    <td>pBAD30</td>
+
                                    <td>concentration of recycled captured bacteria </td>
−
                                    <td>Detecting effectiveness of protein LBT5</td>
+
 
                                   </tr>
 
                                   </tr>
 +
                          </tbody>
 +
                        </table>
 +
                    </div>
 +
                    <p>The description of parameter, their values and the references involved in this model are listed in the second table.</p>
 +
                    <div class="table-responsive " style="padding: 10px 100px; text-align: center; font-size:14px;">
 +
                        <table class="table table-hover">
 +
                          <thead>
 +
                              <tr>
 +
                                <th style="text-align: center;">Parameter</th>
 +
                                <th style="text-align: center;">Description</th>
 +
                              </tr>
 +
                          </thead>
 +
 +
                          <tbody>
 
                                   <tr>
 
                                   <tr>
−
                                     <td>pmrA-pmrB/LBT6-pmrC-GFP </td>
+
                                     <td>dif</td>
−
                                    <td>pBAD30</td>
+
                                     <td>the diffuse ability of bacteria</td>
−
                                     <td> Detecting effectiveness of protein LBT6</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT7-pmrC-GFP</td>
+
                                    <td>difl</td>
−
                                    <td>pBAD30 </td>
+
                                    <td>the diffuse ability of lanthanide ion</td>
−
                                    <td> Detecting effectiveness of protein LBT7</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT8-pmrC-GFP</td>
+
                                    <td>deadp</td>
−
                                    <td>pBAD30 </td>
+
                                    <td>dead/birth per minute of bacteria</td>
−
                                    <td>Detecting effectiveness of protein LBT8</td>
+
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT9-pmrC-GFP</td>
+
                                    <td>vlcx</td>
−
                                    <td> pBAD30 </td>
+
                                    <td>velocity of movement on x label</td>
−
                                    <td>Detecting effectiveness of protein LBT9</td>
+
 
                                   </tr>
 
                                   </tr>
−
                                  <tr>
+
 
−
                                    <td>pmrA-pmrB/LBT10-pmrC-GFP</td>
+
                                  <tr>
−
                                    <td>pBAD30 </td>
+
                                    <td> vlcy </td>
−
                                    <td> Detecting effectiveness of protein LBT10</td>
+
                                    <td>velocity of movement on z label</td>
 
                                   </tr>
 
                                   </tr>
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT11-pmrC-GFP</td>
+
                                    <td>vlcz </td>
−
                                    <td>pBAD30 </td>
+
                                    <td>concentration of recycled captured bacteria </td>
−
                                    <td>Detecting effectiveness of protein LBT11</td>
+
 
                                   </tr>
 
                                   </tr>
 +
 
                                   <tr>
 
                                   <tr>
−
                                    <td>pmrA-pmrB/LBT12-pmrC-GFP </td>
+
                                    <td>krec </td>
−
                                    <td> pBAD30 </td>
+
                                    <td> speed of bacteria recycled on Si-board </td>
−
                                    <td>Detecting effectiveness of protein LBT12</td>
+
                                  </tr>
 +
                                  <tr>
 +
                                    <td>kabcs</td>
 +
                                    <td>speed of bacteria recycled on Si-board</td>
 +
                                  </tr>
 +
 
 +
                                  <tr>
 +
                                    <td> rmax </td>
 +
                                    <td> the amount of bacteria recyclyed on each area of Si-board </td>
 +
                                  </tr>
 +
                                  <tr>
 +
                                    <td>csen</td>
 +
                                    <td>the lowest concentration lanthanide ion that can be sensed</td>
 +
                                  </tr>
 +
 
 +
                                  <tr>
 +
                                    <td>ksen </td>
 +
                                    <td>speed of bacteria sensing the lanthanide ion  </td>
 +
                                  </tr>
 +
                                  <tr>
 +
                                    <td>kcap </td>
 +
                                    <td> speed of sensed bacteria capturing the lanthanide ion  </td>
 +
                                  </tr>
 +
 
 +
                                  <tr>
 +
                                    <td>ccap </td>
 +
                                    <td>the lowest concentration lanthanide ion that can be captured </td>
 +
                                  </tr>
 +
                                  <tr>
 +
                                    <td>LBlever</td>
 +
                                    <td> The amount of lanthanide ion captured by one bacteria</td>
 
                                   </tr>
 
                                   </tr>
−
                               
 
 
                           </tbody>
 
                           </tbody>
 
                         </table>
 
                         </table>
Line 529: Line 563:
  
  
−
                 <div id="section2" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">  
+
                 <div id="section4" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">        
−
                     <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Recycling part </h3>
+
                     <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Basical Diffusion</h3>  
−
                     <p>
+
                     <p>Rome was not built in a day. At the beginning, we built a basical diffusion model, just setting the diffusion function in our model.</p>
−
                            The recycling part is to capture the rare earth ions in the environment and to anchor our cells over the silicon net.
+
                    <p>The function is:</p>
−
                     </p>
+
                    <img class="col-xs-3 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/5/58/2017_HUST_China_model_equation1.png" />
−
                     <h4><strong>①With appropriate primers, PCR was carried out to amplify target gene oprf-LBT1-12 and oprf-Si tag.</strong> </h4>
+
 
−
                     <div class="container experiment_circuit col-md-6 col-xs-6 col-md-offset-3 col-xs-offset-3">
+
                    <p class="col-sm-12">We release some Reebot to the top of our hypothetical container as initial value of f<sub>1<sub>, and use ‘ode’ tool in MATLAB to run this function. Here is the result:</p>
−
                         <img src="https://static.igem.org/mediawiki/2017/7/73/2017_HUST_China_Experiment_image2.png" alt="experiment_circuit" class="img-responsive">
+
 
 +
                    <section class="col-xs-6 col-xs-offset-1">
 +
                        <figure>
 +
                            <video controls="controls" max-width="600" height="480" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
 +
                                <source type="video/webm" src="https://static.igem.org/mediawiki/2017/f/fb/2017_HUST_China_model_vedio1.1.webm" />
 +
                                <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
 +
                                <track src="subs/TOS-japanese.srt" kind="subtitles" srclang="jp" label="Japanese" />
 +
                                <track src="subs/TOS-english.srt" kind="subtitles" srclang="en" label="English" />
 +
                                <track src="subs/TOS-turkish.srt" kind="subtitles" srclang="tr" label="Turkish" />
 +
                                <track src="subs/TOS-ukrainian.srt" kind="subtitles" srclang="uk" label="Ukrainian" />
 +
                            </video>
 +
                        </figure>
 +
                    </section>
 +
 
 +
                </div>     
 +
                <div id="section5" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;"> 
 +
                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Recycling</h3>
 +
                    <p>Then we let our model become more complex and more powerful. We decided using Si-board to recycle the Reebot, since Reebot have Sitag, a protein tag that can combine with Si, once it was sensed. We insert recycling function in our model.</p>
 +
                     <p>Firstly, we should change the diffusion function:</p>
 +
                     <img class="col-xs-4 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/b/b2/2017_HUST_China_model_equation2.png" />
 +
                    <p class="col-xs-12" style="text-align: center;">(In the end of all function, the ∂f/∂t will be delta.)</p> 
 +
                    <p class="col-xs-12">And we set the area of Si-board, if it is in Si-board area:</p>
 +
                    <img class="col-sm-5 col-sm-offset-4" src="https://static.igem.org/mediawiki/2017/5/5e/2017_HUST_China_model_equation7.png" />
 +
                    <p class="col-sm-12">Bacteria that combined with Si-board maybe abscess. So taking the abscission into consideration, we should add this function:</p>
 +
                    <img class="col-xs-4 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/1/14/2017_HUST_China_model_equation3.png" />
 +
                     <p class="col-xs-12">Finally:</p>
 +
                    <img class="col-xs-3 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/9/92/2017_HUST_China_model_equation4.png" />
 +
                    <p class="col-sm-12">Set a 5×5 Si-board on the top and then run the model, the result is:</p>
 +
                    <section class="col-xs-6 col-xs-offset-1">
 +
                        <figure>
 +
                            <video controls="controls" width="600" height="480" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
 +
                                <source type="video/webm" src="https://static.igem.org/mediawiki/2017/a/a5/2017_HUST_China_model_vedio2.1.webm" />
 +
                                <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
 +
                                <track src="subs/TOS-japanese.srt" kind="subtitles" srclang="jp" label="Japanese" />
 +
                                <track src="subs/TOS-english.srt" kind="subtitles" srclang="en" label="English" />
 +
                                <track src="subs/TOS-turkish.srt" kind="subtitles" srclang="tr" label="Turkish" />
 +
                                <track src="subs/TOS-ukrainian.srt" kind="subtitles" srclang="uk" label="Ukrainian" />
 +
                            </video>
 +
                        </figure>
 +
                    </section> 
 +
 
 +
                </div>
 +
                <div id="section6" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">
 +
                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Sensing and Capture</h3>
 +
                    <p>This is the biggest part of our model. We should take lanthanide ion, sensing and capture into consideration. So, our data doesn’t just contain concentration of bacteria anymore. It now contains four: concentration of initially bacteria (haven’t sensed the lanthanide ion) as f<sub>1</sub>, concentration of lanthanide ion as f<sub>2</sub>, concentration of sensed bacteria（sensed the lanthanide ion but haven’t captured）as f<sub>3</sub> and concentration of captured bacteria (have captured enough lanthanide ion) as f<sub>4</sub>.</p>
 +
                    <img class="col-xs-4 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/5/5b/2017_HUST_China_model_equation5.png" />
 +
                    <p class="col-xs-12">And the sensed bacteria will capture the lanthanide ion around it, to simplify the function, we just consider it at two status: no captured and full captured. The function is:</p>
 +
                    <img class="col-xs-4 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/a/a0/2017_HUST_China_model_equation6.png" />
 +
                    <p class="col-xs-12">We should also modify the recycling function. It will only use to sensed bacteria and captured bacteria. </p>
 +
                    <p class="col-xs-12">Finally, the model is over. Running the model, the result is:</p>
 +
                    <section class="col-xs-6 col-xs-offset-1">
 +
                        <figure>
 +
                            <video controls="controls" width="600" height="480" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
 +
                                <source type="video/webm" src="https://static.igem.org/mediawiki/2017/6/60/2017_HUST_China_model_vedio3.2.webm" />
 +
                                <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
 +
                                <track src="subs/TOS-japanese.srt" kind="subtitles" srclang="jp" label="Japanese" />
 +
                                <track src="subs/TOS-english.srt" kind="subtitles" srclang="en" label="English" />
 +
                                <track src="subs/TOS-turkish.srt" kind="subtitles" srclang="tr" label="Turkish" />
 +
                                <track src="subs/TOS-ukrainian.srt" kind="subtitles" srclang="uk" label="Ukrainian" />
 +
                            </video>
 +
                        </figure>
 +
                    </section>
 +
                    <p class="col-xs-12">For more intuitive, we draw a figure :</p>
 +
                    <p><strong><span class="col-xs-12">Click on the picture and you can see a clearer picture.Then click on the return key of the browser and you can return.</span></strong></p><br/>
 +
                    <a href="https://static.igem.org/mediawiki/2017/3/39/2017_HUST_China_model_image_part.png" rel="lightbox-demo" title="my caption"><br />
 +
                         <img title="demo1" src="https://static.igem.org/mediawiki/2017/3/39/2017_HUST_China_model_image_part.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 +
                    </a>
 +
                    <p class="col-xs-12">Put these curve into one figure:</p>
 +
                    <p><strong><span class="col-xs-12">Click on the picture and you can see a clearer picture.Then click on the return key of the browser and you can return.</span></strong></p><br/>
 +
                    <a href="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" rel="lightbox-demo" title="my caption"><br />
 +
                        <img title="demo1" src="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 +
                    </a>
 +
                    <p class="col-xs-12">It’s clearly that there are three stages during the whole simulation. The first stage is sensing, second stage is capturing and third is recycling.</p>
 +
                    <div class="col-xs-12">
 +
                        <h4 style="color:#97477c"><strong>Result:</strong></h4>
 +
                        <p>Firstly, we run a model which is: there are enough concentration of lanthanide ion in the consider, and we put some Reebot on the top. Here is the video:</p>
 +
                        <section class="col-xs-6 col-xs-offset-1">
 +
                            <figure>
 +
                                <video controls="controls" width="600" height="480" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
 +
                                    <source type="video/webm" src="https://static.igem.org/mediawiki/2017/6/60/2017_HUST_China_model_vedio3.2.webm" />
 +
                                    <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
 +
                                    <track src="subs/TOS-japanese.srt" kind="subtitles" srclang="jp" label="Japanese" />
 +
                                    <track src="subs/TOS-english.srt" kind="subtitles" srclang="en" label="English" />
 +
                                    <track src="subs/TOS-turkish.srt" kind="subtitles" srclang="tr" label="Turkish" />
 +
                                    <track src="subs/TOS-ukrainian.srt" kind="subtitles" srclang="uk" label="Ukrainian" />
 +
                                </video>
 +
                            </figure>
 +
                        </section>
 
                     </div>
 
                     </div>
−
                     <h4 class="col-xs-12"><strong>②Then we constructed two vectors to achieve expression of LBT and Si-tag.</strong> </h4>
+
                     <div class="col-xs-12">
−
                    <div class="container experiment_circuit col-md-6 col-xs-6 col-md-offset-3 col-xs-offset-3">
+
                        <p> From the video, we can see that once we put Reebot into the container, the Reebot quickly diffuse. At the same time, Reebot sense the lanthanide ion around them, and express more LBT (lanthanide binding tag), sitag on their surface. Then, the LBT start to capture the lanthanide ion in the environment. As time goes by, Reebot have captured more and more lanthanide. At the beginning, the concentration of lanthanide ion is 0.02，and now, it is 0.006.</p>
−
                        <img src="https://static.igem.org/mediawiki/2017/7/78/2017_HUST_China_Experiment_image3.png" alt="experiment_circuit" class="img-responsive">
+
                        <p>When t = 60, we set a 6*8 Si-board on the top. The sensed bacteria and captured bacteria are recycling to the Si-board. You can see it clearly that the edge of Si-board recycled more Reebot. In the end, many bacteria are recycled to the Si-board, but there are still many Reebot in the environment. The reason is concentration gradient caused by Si-board isn’t large enough so that the diffusion isn’t effective. But keeping the Si-board here doesn’t cost much, so we can stand it.</p>
 +
                        <p>In order to show you more details, we sum each areas’ concentration and put them into on figure:</p>
 +
                        <p><strong><span class="col-xs-12">Click on the picture and you can see a clearer picture.Then click on the return key of the browser and you can return.</span></strong></p><br/>
 +
                        <a href="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" rel="lightbox-demo" title="my caption"><br />
 +
                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 +
                        </a>
 
                     </div>
 
                     </div>
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                     <p class="col-xs-12">
+
                     <div class="col-xs-12">
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                         We cloned the sequence of oprf-LBT and Si-tag into the vector pET28α in order to make the sequence be tagged N-terminally with a 6×His tag. 6×His tag is an affinity tag which allows the tagged recombinant protein to be purified in the process of affinity purification.  
+
                         <p> From the figure, we can see lot of things. In the first 60 units of time, most Reebot quickly captured enough bacteria, the concentration of initial bacteria and sensed bacteria are very low. Then, the Si-board set, Reebot begin to recycle. At first, in the figure, Reebot recycle at about a same speed. Then recycle speed is slower and slower. </p>
−
                    </p>
+
                        <p>This model simulate the ordinary use of Reebot. the result can show us the detail of process, we can determine how much Reebot to release and recycle for how long upon that.</p>
−
                    <h4><strong>③Detection</strong> </h4>
+
                        <p>Then, we run an other simulation to show the intelligence of Reebot. In this simulation, there was no lanthanide ion at the beginning, we put lanthanide ion at some certain time.</p>
−
                    <p>
+
                        <p>The video and figure is here.</p>
−
                      In the meanwhile,we added the FLAG tag to achieve that whether the LBTs and the Si tag was expressed on the cell membrane.  
+
                        <section class="col-xs-6 col-xs-offset-1">
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                    </p>
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                            <figure>
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                </div>
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                                <video controls="controls" width="600" height="480" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
−
 
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                                    <source type="video/webm" src="https://2017.igem.org/File:2017_HUST_China_model_vedio4.2.webm" />
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                <div id="section3" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">        
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                                    <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
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                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Construction of the whole circuit</h3>
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                    <p>
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                                    <track src="subs/TOS-english.srt" kind="subtitles" srclang="en" label="English" />
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                      All of our part should be built in one plasmid. Therefore, we use endonuclease and ligase to  replace the GFP by Oprf-LBT and Oprf-Si tag.
+
                                    <track src="subs/TOS-turkish.srt" kind="subtitles" srclang="tr" label="Turkish" />
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                    </p>
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                    <div class="container experiment_circuit col-md-6 col-xs-6 col-md-offset-3 col-xs-offset-3" style="padding: 5px;">
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                                </video>
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                        <img src="https://static.igem.org/mediawiki/2017/5/58/2017_HUST_China_Experiment_image4.png" alt="experiment_circuit" class="img-responsive">
+
                            </figure>
 +
                        </section>
 +
                    </div>
 +
                    <div class="col-xs-12">
 +
                        <p>At the beginning, we release Reebot on the top. While there is no lanthanide ion in environment, so Reebot keep the initial status, and diffuse in the container. When passing through 40 units of time, there are some lanthanide ion release at the bottom of container. From the video, Reebot take a quick reaction, capturing the lanthanide ion and recycling. When it passes 180 units of time, other lanthanide ion release at another position in bottom. Reebot successfully capture them too.</p>
 +
                        <p>It’s also our project’s future plan. Once we achieve it, we can release Reebot in factory’s pool. If there is no lanthanide ion, Reebot just live as a normal escherichia coli. And once lanthanide ion emerge, Reebot can sense and capture it, then Reebot combine with Si-board. </p>
 
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            <h3><strong>Acknowledgments:</strong></h3>
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Revision as of 13:52, 28 October 2017

Attributions

「Attributions」

Overview

We build an ecosystem level model, in order to see how the Reebot diffuse, sense and capture the lanthanide ion, and be recycled on Si-board. Then we can determine how should we release Reebot, how to recycle the Reebot and how much Reebot do we need to produce the best result. The ecosystem could derive the best strategy to make full use of Reebot.

Assumptions

The variability of temperature has no effect on our Reebot or it could be ignored.

Liquid in the container is stationary.

There is nothing else in the container which can influence Reebot.

Constant & Parameter

The description of constant, their values and the references involved in this model are listed in this table.

Constant Description
f1 concentration of Initially Bacteria
f2 concentration of Lanthanide ion
f3 concentration of Sensed Bacteria
f4 concentration of Captured Bacteria
f5 concentration of recycled sensed bacteria
f6 concentration of recycled captured bacteria

The description of parameter, their values and the references involved in this model are listed in the second table.

Parameter Description
dif the diffuse ability of bacteria
difl the diffuse ability of lanthanide ion
deadp dead/birth per minute of bacteria
vlcx velocity of movement on x label
vlcy velocity of movement on z label
vlcz concentration of recycled captured bacteria
krec speed of bacteria recycled on Si-board
kabcs speed of bacteria recycled on Si-board
rmax the amount of bacteria recyclyed on each area of Si-board
csen the lowest concentration lanthanide ion that can be sensed
ksen speed of bacteria sensing the lanthanide ion
kcap speed of sensed bacteria capturing the lanthanide ion
ccap the lowest concentration lanthanide ion that can be captured
LBlever The amount of lanthanide ion captured by one bacteria

Basical Diffusion

Rome was not built in a day. At the beginning, we built a basical diffusion model, just setting the diffusion function in our model.

The function is:

We release some Reebot to the top of our hypothetical container as initial value of f1, and use ‘ode’ tool in MATLAB to run this function. Here is the result:

Recycling

Then we let our model become more complex and more powerful. We decided using Si-board to recycle the Reebot, since Reebot have Sitag, a protein tag that can combine with Si, once it was sensed. We insert recycling function in our model.

Firstly, we should change the diffusion function:

(In the end of all function, the ∂f/∂t will be delta.)

And we set the area of Si-board, if it is in Si-board area:

Bacteria that combined with Si-board maybe abscess. So taking the abscission into consideration, we should add this function:

Finally:

Set a 5×5 Si-board on the top and then run the model, the result is:

Sensing and Capture

This is the biggest part of our model. We should take lanthanide ion, sensing and capture into consideration. So, our data doesn’t just contain concentration of bacteria anymore. It now contains four: concentration of initially bacteria (haven’t sensed the lanthanide ion) as f1, concentration of lanthanide ion as f2, concentration of sensed bacteria（sensed the lanthanide ion but haven’t captured）as f3 and concentration of captured bacteria (have captured enough lanthanide ion) as f4.

And the sensed bacteria will capture the lanthanide ion around it, to simplify the function, we just consider it at two status: no captured and full captured. The function is:

We should also modify the recycling function. It will only use to sensed bacteria and captured bacteria.

Finally, the model is over. Running the model, the result is:

For more intuitive, we draw a figure :

Click on the picture and you can see a clearer picture.Then click on the return key of the browser and you can return.



demo1

Put these curve into one figure:

Click on the picture and you can see a clearer picture.Then click on the return key of the browser and you can return.



demo1

It’s clearly that there are three stages during the whole simulation. The first stage is sensing, second stage is capturing and third is recycling.

Result:

Firstly, we run a model which is: there are enough concentration of lanthanide ion in the consider, and we put some Reebot on the top. Here is the video:

From the video, we can see that once we put Reebot into the container, the Reebot quickly diffuse. At the same time, Reebot sense the lanthanide ion around them, and express more LBT (lanthanide binding tag), sitag on their surface. Then, the LBT start to capture the lanthanide ion in the environment. As time goes by, Reebot have captured more and more lanthanide. At the beginning, the concentration of lanthanide ion is 0.02，and now, it is 0.006.

When t = 60, we set a 6*8 Si-board on the top. The sensed bacteria and captured bacteria are recycling to the Si-board. You can see it clearly that the edge of Si-board recycled more Reebot. In the end, many bacteria are recycled to the Si-board, but there are still many Reebot in the environment. The reason is concentration gradient caused by Si-board isn’t large enough so that the diffusion isn’t effective. But keeping the Si-board here doesn’t cost much, so we can stand it.

In order to show you more details, we sum each areas’ concentration and put them into on figure:

Click on the picture and you can see a clearer picture.Then click on the return key of the browser and you can return.



demo1

From the figure, we can see lot of things. In the first 60 units of time, most Reebot quickly captured enough bacteria, the concentration of initial bacteria and sensed bacteria are very low. Then, the Si-board set, Reebot begin to recycle. At first, in the figure, Reebot recycle at about a same speed. Then recycle speed is slower and slower.

This model simulate the ordinary use of Reebot. the result can show us the detail of process, we can determine how much Reebot to release and recycle for how long upon that.

Then, we run an other simulation to show the intelligence of Reebot. In this simulation, there was no lanthanide ion at the beginning, we put lanthanide ion at some certain time.

The video and figure is here.

At the beginning, we release Reebot on the top. While there is no lanthanide ion in environment, so Reebot keep the initial status, and diffuse in the container. When passing through 40 units of time, there are some lanthanide ion release at the bottom of container. From the video, Reebot take a quick reaction, capturing the lanthanide ion and recycling. When it passes 180 units of time, other lanthanide ion release at another position in bottom. Reebot successfully capture them too.

It’s also our project’s future plan. Once we achieve it, we can release Reebot in factory’s pool. If there is no lanthanide ion, Reebot just live as a normal escherichia coli. And once lanthanide ion emerge, Reebot can sense and capture it, then Reebot combine with Si-board.

Acknowledgments: