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

 
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             <h1 class="tab-h1" ><strong>「Interlab」</strong></h1>
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             <h1 class="tab-h1" ><strong>「Model」</strong></h1>
 
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                     <li class="active"><a href="#section1">Introduction</a></li>
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                     <li class="active"><a href="#section1">Overview</a></li>
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                     <li><a href="#section2">Provenance and Release</a></li>
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                     <li><a href="#section2">Assumptions</a></li>
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                     <li><a href="#section3">Chassis and Safety </a></li>
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                     <li><a href="#section3">Constant & Parameter</a></li>
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                     <li><a href="#section4">Instrument</a></li>
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                     <li><a href="#section4">Basical Diffusion</a></li>
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                     <li><a href="#section5">Calibration Protocol</a></li>  
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                     <li><a href="#section5">Recycle</a></li>
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                     <li><a href="#section6">Cell Culture</a></li>  
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                     <li><a href="#section6">Sense and Enrichment</a></li>
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                     <li><a href="#section7">Interlab Result</a></li>  
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                     <li><a href="#section7">Result</a></li>
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                 <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;">Introduction</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 style="padding:10px">It is significant for synthetic biology to develop a reliable and repeatable measurement, the same to all the other engineering disciplines. We HUST-China have volunteered to test some RBS devices (BCDs) that are intended to make gene expression more precise and reliable by measure the expression level of GFP, in order to help the iGEM community collect data about how reliable will these devices turn out to be in labs around the world.
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                     <p>We build an ecosystem level model, in order to see how the Reebot diffuse, sense and enrich the lanthanide ions, 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 for the best result. The ecosystem could derive the best strategy to make full use of Reebot.
 
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                 <div id="section2" 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;">Provenance and Release</h3>
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                     <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Assumptions</h3>
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                    <h4><strong>①Individuals responsible for conducting InterLab study</strong> </h4>
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                    <p>The variability of temperature has no effect on our Reebot or it could be ignored.</p>
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                    <p>Liquid in the container is stationary.</p>
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                    <p>There is nothing else in the container which can influence Reebot.</p>  
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                </div>
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                <div id="section3" 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;">Constant & Parameter</h3>
 +
                    <p>The description of constant, their values and the references involved in this model are listed in this table.</p>
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                                 <th style="text-align: center;">Individuals</th>
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                                 <th style="text-align: center;">Constant </th>
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                                 <th style="text-align: center;">Interlab Part</th>
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                                <td>Kangyuan Yu, Haibo Huang, Ziyang Xiao, Shaofeng Liao</td>
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                                    <td>f<sub>1</sub></td>
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                                <td>created the devices</td>
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                                    <td>concentration of Initially Bacteria</td>
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                              </tr>
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                                <td>Efan Wang, Long Cheng, HuiPing Shi </td>
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                                    <td>f<sub>2</sub></td>
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                                <td>conducted the measurements</td>
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                                    <td>concentration of Lanthanide ion</td>
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                              </tr>
+
                                  </tr>
−
                              <tr>
+
                                  <tr>
−
                                <td>Efan Wang</td>
+
                                    <td>f<sub>3</sub></td>
−
                                <td>processed the data</td>
+
                                    <td>concentration of Sensed Bacteria</td>
−
                              </tr>
+
                                  </tr>
 +
                                  <tr>
 +
                                    <td>f<sub>4</sub></td>
 +
                                    <td>concentration of Recycled Bacteria</td>
 +
                                  </tr>
 +
                                  <tr>
 +
                                    <td>f<sub>5</sub></td>
 +
                                    <td>concentration of recycled sensed bacteria </td>
 +
                                  </tr>
 +
                                  <tr>
 +
                                    <td>f<sub>6</sub></td>
 +
                                    <td>concentration of enriched recycled bacteria </td>
 +
                                  </tr>
 
                           </tbody>
 
                           </tbody>
 
                         </table>
 
                         </table>
 
                     </div>
 
                     </div>
−
                     <h4><strong> ②Corresponding email</strong> </h4>
+
                     <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;">
+
                    <div class="table-responsive " style="padding: 10px 100px; text-align: center; font-size:14px;">
 
                         <table class="table table-hover">
 
                         <table class="table table-hover">
 
                           <thead>
 
                           <thead>
 
                               <tr>
 
                               <tr>
−
                                 <th style="text-align: center;">Individuals </th>
+
                                 <th style="text-align: center;">Parameter</th>
−
                                 <th style="text-align: center;">Emails</th>
+
                                 <th style="text-align: center;">Description</th>
 
                               </tr>
 
                               </tr>
 
                           </thead>
 
                           </thead>
 +
 
                           <tbody>
 
                           <tbody>
−
                              <tr>
+
                                  <tr>
−
                                <td>Efan Wang</td>
+
                                    <td>dif</td>
−
                                <td>erfan@hust.edu.cn</td>
+
                                    <td>the diffuse ability of bacteria</td>
−
                              </tr>
+
                                  </tr>
−
                              <tr>
+
                                  <tr>
−
                                <td>Kangyuan Yu</td>
+
                                    <td>difl</td>
−
                                <td>985930862@qq.com</td>
+
                                    <td>the diffuse ability of lanthanide ion</td>
−
                              </tr>
+
                                  </tr>
−
                              <tr>
+
                                  <tr>
−
                                <td>Haibo Huang </td>
+
                                    <td>deadp</td>
−
                                <td>u201512127@hust.edu.cn</td>
+
                                    <td>dead/birth per minute of bacteria</td>
−
                              </tr>
+
                                  </tr>
−
                              <tr>
+
                                  <tr>
−
                                <td>Ziyang Xiao</td>
+
                                    <td>vlcx</td>
−
                                <td>372657289@qq.com</td>
+
                                    <td>velocity of movement on x label</td>
−
                              </tr>
+
                                  </tr>
−
                              <tr>
+
 
−
                                <td>Shaofeng Liao</td>
+
                                  <tr>
−
                                <td> 15827233830@qq.com</td>
+
                                    <td> vlcy </td>
−
                              </tr>
+
                                    <td>velocity of movement on z label</td>
−
                              <tr>
+
                                  </tr>
−
                                <td>HuiPing Shi</td>
+
                                  <tr>
−
                                <td>172295915@qq.com</td>
+
                                    <td>vlcz </td>
−
                              </tr>
+
                                    <td>concentration of enriched recycle bacteria </td>
−
                              <tr>
+
                                  </tr>
−
                                <td>Long Cheng</td>
+
 
−
                                <td>u201512127@hust.edu.cn</td>
+
                                  <tr>
−
                              </tr>
+
                                    <td>krec </td>
 +
                                    <td> speed of bacteria recycled on Si-board </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 enriched </td>
 +
                                  </tr>
 +
                                  <tr>
 +
                                    <td>LBlever</td>
 +
                                    <td> The amount of lanthanide ion enriched by one bacteria</td>
 +
                                  </tr>
 
                           </tbody>
 
                           </tbody>
 
                         </table>
 
                         </table>
 
                     </div>
 
                     </div>
−
                </div>       
 
−
                <div id="section3" 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;">Chassis and Safety</h3>
 
−
                    <h4><strong> What chassis did you use?</strong> </h4>
 
−
                    <p>Escherichia coli DH5alpha</p>
 
−
                    <h4><strong> What Biosafety Level is your chassis? </strong> </h4>
 
−
                    <p>BSL1</p>
 
−
                    <h4><strong>What PPE did you utilize during your experiments? </strong></h4>
 
−
                    <p>Tianming gloves</p>
 
−
                    <p>Songxinjiujiu labcoats</p>
 
 
                 </div>
 
                 </div>
−
                <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;">Instrument</h3>
 
−
                    <h4><strong> What instrument did you use during your measurements? </strong> </h4>
 
−
                    <p>plate reader</p>
 
−
                    <h4><strong> Please provide the brand and model of your instrument.</strong> </h4>
 
−
                    <p>Flexstation 3</p>
 
−
                </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;">Calibration Protocol</h3>
 
  
  
−
                    <h4><strong>  A1. Protocol for Optical Density (OD600) Standard Measurement</strong> </h4>
+
                <div id="section4" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">        
−
                     <h5><strong>Did you use pathlength correction during measurement? </strong> </h5>
+
                     <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Basical Diffusion</h3>   
−
                    <p>Yes</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>
−
                    <h5><strong>Number of flashes per well  </strong> </h5>
+
                     <p>The function is:</p>
−
                    <p>6</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" />
−
                    <h5><strong>Orbital averaging (mm)   </strong> </h5>
+
−
                     <p>600</p>
+
−
                    <h5><strong> What temperature setting did you use during the measurement? </strong> </h5>
+
−
                     <p>22℃</p>
+
−
                     <h5><strong>What type of 96-well plate did you use? </strong> </h5>
+
−
                    <p>Black plate (preferred)</p>
+
−
                    <h5><strong> Did your plate have flat-bottomed or round-bottomed wells? </strong> </h5>
+
−
                    <p>Flat</p>
+
  
 +
                    <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>
  
−
                     <h4><strong> A2. Measurement Steps</strong></h4>
+
                     <section class="col-xs-6 col-xs-offset-2">
−
                                <li class="dropdown" style="list-style-type:none;">
+
                        <figure>
−
                                    <button href="#" class="dropdown-toggle" data-toggle="dropdown">
+
                            <video controls max-width="400" height="300" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
−
                                        More details <b class="caret"></b>
+
                                <source type="video/webm" src="https://static.igem.org/mediawiki/2017/f/fb/2017_HUST_China_model_vedio1.1.webm" />
−
                                    </button>
+
                                <source type="video/mp4" src="https://static.igem.org/mediawiki/2017/e/ea/HUST-modeling1.mp4" />
−
                                    <ul class="dropdown-menu container" style="padding: 10px;background:#ccc;">
+
                            </video>
−
                                        <li>① Prepare your 96 well plate</li>
+
                        </figure>
−
                                        <li>②Add 100 μl LUDOX 100 % into wells A1, B1, C1, D1</li>
+
                    </section>
−
                                        <li>③Add 100 μl of dH2O into A2, B2, C2, D2</li>
+
−
                                        <li>④Measure absorbance 600 nm of all samples in all standard measurement modes in instrument</li>
+
−
                                        <li>⑤Import data into "Abs600" blue cells in provided Excel calibration sheet</li>
+
−
                                    </ul>
+
−
                                </li>
+
  
−
                   
+
                </div>    
−
                    <h4><strong>  B1. Protocol for FIuorescein Fluoresence standard curve</strong> </h4>
+
                <div id="section5" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">   
−
                     <h5><strong>Did you use pathlength correction during measurement? </strong> </h5>
+
                     <h3 class="col-xs-12" style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Recycle</h3>
−
                     <p>Yes</p>
+
                     <p class="col-xs-12">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 recycle function in our model.</p>
−
                    <h5><strong>Number of flashes per well  </strong> </h5>
+
                     <p class="col-xs-12">Firstly, we should change the diffusion function:</p>
−
                     <p>6</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" />
−
                    <h5><strong>What gain setting did you use?  </strong> </h5>
+
                     <p class="col-xs-12" style="text-align: center;">(In the end of all function, the ∂f/∂t will be delta.)</p>
−
                     <p>Automatic</p>
+
                     <p class="col-xs-12">And we set the area of Si-board, if it is in Si-board area:</p>
−
                     <h5><strong> If you used a filter, what light wavelengths did it pass?  </strong> </h5>
+
                    <img class="col-xs-5 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/5/5e/2017_HUST_China_model_equation7.png" />
−
                     <p>530nm</p>
+
                     <p class="col-xs-12">Bacteria that combined with Si-board maybe abscess. So taking the abscission into consideration, we should add this function:</p>
−
                     <h5><strong>Emission wavelength </strong> </h5>
+
                     <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>530nm</p>
+
                     <p class="col-xs-12">Finally:</p>  
−
                     <h5><strong> Excitation wavelength </strong> </h5>
+
                     <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>485nm</p>
+
                     <p class="col-sm-12">Set a 5×5 Si-board on the top and then run the model, the result is:</p>
−
                     <h5><strong> Fluorescence reading  </strong> </h5>
+
                     <section class="col-xs-6 col-xs-offset-2">
−
                    <p>Bottom optic</p>
+
                        <figure>
−
                    <h5><strong>What type of 96-well plate did you use? </strong> </h5>
+
                            <video controls width="400" height="300" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
−
                    <p>Black plate (preferred)</p>
+
                                <source type="video/webm" src="https://static.igem.org/mediawiki/2017/a/a5/2017_HUST_China_model_vedio2.1.webm" />
−
                    <h5><strong> Did your plate have flat-bottomed or round-bottomed wells? </strong> </h5>
+
                                <source type="video/mp4" src="https://static.igem.org/mediawiki/2017/e/ea/HUST-modeling1.mp4" />
−
                    <p>Flat</p>
+
                                <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
−
                    <h5><strong> What temperature setting did you use during the measurement?  </strong> </h5>
+
                                <track src="subs/TOS-japanese.srt" kind="subtitles" srclang="jp" label="Japanese" />
−
                     <p>22℃</p>
+
                                <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>  
  
−
                    <h4><strong> B2. Measurement Steps</strong></h4>
+
                </div>
−
                     <h5><strong> Part 1: Prepare the Fluorescein stock solution </strong> </h5>
+
                <div id="section6" style="border: solid 1px #666; margin:5px;border-radius:10px;overflow: hidden;">  
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                            <li class="dropdown" style="list-style-type:none;">
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                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px; margin-top:0;">Sense and Enrichment</h3>
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                                <button href="#" class="dropdown-toggle" data-toggle="dropdown">
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                     <p>This is the biggest part of our model. We should take lanthanide ion, sensing and enrichment 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 recycled）as f<sub>3</sub> and concentration of recycled bacteria (have enriched enough lanthanide ion) as f<sub>4</sub>.</p>
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                                    More details <b class="caret"></b>
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                    <img class="col-xs-4 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/5/5b/2017_HUST_China_model_equation5.png" />
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                                </button>
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                    <p class="col-xs-12">And the sensed bacteria will enrich the lanthanide ion around it, to simplify the function, we just consider it at two status: no enriched and full enriched. The function is:</p>
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                                <ul class="dropdown-menu" style="padding: 10px; background:#ccc;">
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                    <img class="col-xs-4 col-xs-offset-4" src="https://static.igem.org/mediawiki/2017/a/a0/2017_HUST_China_model_equation6.png" />
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                                    <li>①  Spin down fluorescein stock tube to make sure pellet is at the bottom of tube.</li>
+
                    <p class="col-xs-12">We should also modify the enrichment function. It will only use to sensed bacteria and enriched bacteria. </p>
−
                                    <li>②  Prepare 2x fluorescein stock solution (100 μM) by resuspending fluorescein in 1ml of 1xPBS</li>
+
                    <p class="col-xs-12">Finally, the model is over. Running the model, the result is:</p>
−
                                    <li>③  Dilute the 2x fluorescein stock solution using 1xPBS to make a 1x fluorescein solution (final concentration is 50 μM).</li>
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                    <section class="col-xs-6 col-xs-offset-2">
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                                    <li>④  *Illustration of serial dilution samples in 96 well plate or cuvettes: value decreases by 2-fold with each column (50% in column 2, 25% in column 3, 12.5% in column 4, etc.)</li>
+
                        <figure>
−
                                    <li>⑤Import data into "Abs600" blue cells in provided Excel calibration sheet</li>
+
                            <video controls width="400" height="300" 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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                                 </ul>
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                                <source type="video/webm" src="https://static.igem.org/mediawiki/2017/6/60/2017_HUST_China_model_vedio3.2.webm" />
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                             </li>
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                                <source type="video/mp4" src="https://static.igem.org/mediawiki/2017/e/ea/HUST-modeling3.mp4" />
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                     <h5>Measurement work flow：</h5>
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                                <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/>
 
                     <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/>
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                     <a href="https://static.igem.org/mediawiki/2017/f/f8/2017_HUST_China_interlab_img_001.png" rel="lightbox-demo" title="my caption"><br />
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                     <a href="https://static.igem.org/mediawiki/2017/3/39/2017_HUST_China_model_image_part.png" rel="lightbox-demo" title="my caption"><br />
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                         <img title="demo1" src="https://static.igem.org/mediawiki/2017/f/f8/2017_HUST_China_interlab_img_001.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
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                         <img src="https://static.igem.org/mediawiki/2017/3/39/2017_HUST_China_model_image_part.png" alt="figure" class="col-xs-8 col-xs-offset-1">
 
                     </a>
 
                     </a>
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                     <div class="col-xs-12" >
+
                     <p class="col-xs-12">Put these curve into one figure:</p>  
−
                        <h5><strong>  Part 2: Prepare the serial dilutions of Fluorescein </strong> </h5>
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                        <li class="dropdown" style="list-style-type:none;">
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                                <button href="#" class="dropdown-toggle" data-toggle="dropdown">
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                                    More details <b class="caret"></b>
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                                </button>
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                                <ul class="dropdown-menu" style="padding: 10px; background:#ccc;">
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                                        <li>①Add 100 μl of 1xPBS into wells A2, B2, C2, D2....A12, B12, C12, D12.</li>
+
−
                                        <li>②Add 200 μl of 1x Fluorescein stock solution into A1, B1, C1, D1.</li>
+
−
                                        <li>③  Transfer 100 μl of Fluorescein stock solution from A1 into A2.</li>
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−
                                        <li>④  Mix A2 by pipetting up and down 3x and transfer 100 μl into A3.</li>
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−
                                        <li>⑤  Mix A3 by pipetting up and down 3x and transfer 100 μl into A4.</li>
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                                        <li>⑥  Mix A4 by pipetting up and down 3x and transfer 100 μl into A5.</li>
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                                        <li>⑦  Mix A5 by pipetting up and down 3x and transfer 100 μl into A6.</li>
+
−
                                        <li>⑧  Mix A6 by pipetting up and down 3x and transfer 100 μl into A7.</li>
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                                        <li>⑨  Mix A7 by pipetting up and down 3x and transfer 100 μl into A8.</li>
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−
                                        <li>⑩  Mix A8 by pipetting up and down 3x and transfer 100 μl into A9.</li>
+
−
                                        <li>⑪  Mix A9 by pipetting up and down 3x and transfer 100 μl into A10.</li>
+
−
                                        <li>⑫  Mix A10 by pipetting up and down 3x and transfer 100 μl into A11.</li>
+
−
                                        <li>⑬  Mix A11 by pipetting up and down 3x and transfer 100 μl into liquid waste.</li>
+
−
                                        <li>⑭  TAKE CARE NOT TO CONTINUE SERIAL DILUTION INTO COLUMN 12.</li>
+
−
                                        <li>⑮  Repeat dilution series for rows B, C, D.
+
−
                                        <li>⑯  Measure fluorescence of all samples in all standard measurement modes in instrument.
+
−
                                </ul>
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                            </li>
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                    </div>
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                </div>
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                <div id="section6" 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;">Cell Culture Setup and Measurement</h3>
+
 
                     <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/>
 
                     <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/e/e6/2017_HUST_China_interlab_img_002.png" rel="lightbox-demo" title="my caption"><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 />
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                         <img title="demo1" src="https://static.igem.org/mediawiki/2017/e/e6/2017_HUST_China_interlab_img_002.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
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                         <img src="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" alt="figure" class="col-xs-8 col-xs-offset-1">
 
                     </a>
 
                     </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 recycle.</p>
 +
                </div>
 +
                <div id="section7" 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;">Result</h3>
 +
                        <h4><strong><I>a. High Concentration Simulation</I></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.And we will set the Si-board later. Here is the video:</p>
 +
                        <section class="col-xs-6 col-xs-offset-2">
 +
                            <figure>
 +
                                <video controls width="400" height="300" 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" />
 +
                                    <source type="video/mp4" src="https://static.igem.org/mediawiki/2017/e/ea/HUST-modeling3.mp4" />
 +
                                    <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 class="col-xs-12">
 
                     <div class="col-xs-12">
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                         <h5><strong>Transformation:</strong> </h5>
+
                         <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 enrich the lanthanide ion in the environment. As time goes by, Reebot have enriched more and more lanthanide. At the beginning, the concentration of lanthanide ion is approximately 0.03 unit，and now, it is about 0.003.</p>
−
                            <ul class="yuandian">
+
                        <p>When t = 50, we set a 6*8 Si-board on the top. The sensed bacteria and enriched bacteria are recycle to the Si-board. You can see it clearly that the edge of Si-board enriched 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. So, in experiment we change the way Si-Board set, like this way.<br /></div><div class="col-xs-12">
−
                                <li> control</li>
+
                                                <a href="https://static.igem.org/mediawiki/2017/d/d8/HUST_recycle_tool.png" rel="lightbox-demo" title="my caption"><br />
−
                                <li> Negative control</li>
+
                            <img  src="https://static.igem.org/mediawiki/2017/d/d8/HUST_recycle_tool.png" alt="figure" class="col-xs-8 col-xs-offset-1">
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                                <li>Test Device 1: J23101+I13504</li>
+
                        </a><div class="col-xs-12">
−
                                <li>Test Device 2: J23106+I13504</li>
+
                     
−
                                <li>Test Device 3: J23117+I13504</li>
+
                        <p><br />But it's quite difficult to simulate this model, so we still use the old container model<br/>
−
                                <li>Test Device 4: J23101.BCD2.E0040.B0015</li>
+
                        In order to show you more details, we sum each areas’ concentration and put them into on figure:</p>
−
                                <li>Test Device 5: J23106.BCD2.E0040.B0015</li>
+
                        <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/>
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                                <li>Test Device 6: J23117.BCD2.E0040.B0015</li>
+
                         <a href="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" rel="lightbox-demo" title="my caption"><br />
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                            </ul>
+
                            <img  src="https://static.igem.org/mediawiki/2017/6/66/2017_HUST_Chian_model_image_all.png" alt="figure" class="col-xs-8 col-xs-offset-1">
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                        <h5><strong>Cell growth:</strong> </h5>
+
                        </a>
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                         <li class="dropdown" style="list-style-type:none;">
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                    </div>
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                                <button href="#" class="dropdown-toggle" data-toggle="dropdown">
+
                    <div class="col-xs-12">
−
                                    More details <b class="caret"></b>
+
                        <p> From the figure, we can see lot of things. In the first 50 units of time, most Reebot quickly enriched enough lanthanide ions, 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 enrich at about a same speed. Then enrichm speed is slower and slower. </p>
−
                                </button>
+
                        <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>
−
                                <ul class="dropdown-menu" style="padding: 10px; background:#ccc;">
+
                    </div>
  
−
                                    <li> ①    Pick 2 colonies from each of plate and inoculate each in 5-10 mL LB medium + Chloramphenicol (For antibiotic concentrations, please follow these guidelines:<a href="http://parts.igem.org/Help:Protocols/Antibiotic_Stocks">http://parts.igem.org/Help:Protocols/Antibiotic_Stocks).
 
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                                    </a>
 
−
                                    </li>
 
−
                                    <li>②  Grow the cells overnight (16-18 hours) at 37°C and 220 rpm.</li>
 
−
                                    <li>③  With 8 total devices (including controls) and 2 colonies per device, you should have 16 cultures.</li>
 
  
−
                                </ul>
+
                    <div class="col-xs-12">
−
                            </li>
+
                         <h4><strong><I>b. Low Concentration Simulation </I></strong></h4>
−
                         <h5><strong>Cell growth, sampling, and assay</strong> </h5>
+
                         <p>According to the paper, the concentration of lanthanide ion is much lower than expected. So in this simulation, we set concentration of lanthanide ion as 0.001 unit. Other parameters are same as before. Here is the result:</p>
−
                         <li class="dropdown" style="list-style-type:none;">
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                        <section class="col-xs-6 col-xs-offset-2">
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                                <button href="#" class="dropdown-toggle" data-toggle="dropdown">
+
                            <figure>
−
                                     More details <b class="caret"></b>
+
                                <video controls width="400" height="300" 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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                                </button>
+
                                    <source type="video/webm" src="https://static.igem.org/mediawiki/2017/4/48/2017_HUST_China_model_vedio6.webm" />
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                                 <ul class="dropdown-menu" style="padding: 10px; background:#ccc;">
+
                                    <source type="video/mp4" src="https://static.igem.org/mediawiki/2017/6/60/HUST-modeling6.mp4" />
 +
                                    <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>
 +
                        <a href="https://static.igem.org/mediawiki/2017/9/9d/2017_HUST_China_model_model2part.png" rel="lightbox-demo" title="my caption"><br />
 +
                            <img  src="https://static.igem.org/mediawiki/2017/9/9d/2017_HUST_China_model_model2part.png" alt="figure" class="col-xs-8 col-xs-offset-1">
 +
                        </a>
 +
                        <a href="https://static.igem.org/mediawiki/2017/f/fe/2017_HUST_China_model_model2.png" rel="lightbox-demo" title="my caption"><br />
 +
                            <img  src="https://static.igem.org/mediawiki/2017/f/fe/2017_HUST_China_model_model2.png" alt="figure" class="col-xs-8 col-xs-offset-1">
 +
                        </a>
 +
                        <p class="col-xs-12">In this situation, Reebot enriched most of lanthanide ion. But actually, some Reebot are wasted. these Reebot sensed lanthanide ion and expressed LBT and Sitag on their surface. But sensed bacteria is so much that only part of them can enrich enough lanthanide ion. Others only enriched a little and be recycled to Si-board.</p>
 +
                    </div>
  
−
                                    <li>①  Set your instrument to read Abs600 (as OD calibration setting)</li>
 
−
                                    <li>②  Measure Abs600 of the overnight cultures</li>
 
−
                                    <li>③  Import data into blue cells in Excel Dilultion Calculation sheets provided</li>
 
−
                                    <li>④  Dilute the cultures to a target Abs600 of 0.02 (see the volume of preloading culture and media in Excel Dilution Calculation sheets) in 12 ml LB medium + Chloramphenicol in 50 mL falcon tube.</li>
 
−
                                    <li>⑤  Remove 500uL of each culture for your zero time point and hold these samples on ice. (You should have 16 sample tubes from the time=0 time point)</li>
 
−
                                    <li>⑥  Incubate the cultures at 37°C and 220 rpm.</li>
 
−
                                    <li>⑦  Remove 500uL samples of each culture after 2, 4 and 6 hours of growth. Keep samples on ice while completing the additional time points. You should have 16 sample tubes for each time point.</li>
 
−
                                    <li>⑧  Set up your four measurement plates: For colony #1 culture, pipet 100 uL samples into wells A, B, C and D of the row for that device. For colony #2, pipet 100uL samples into wells E, F, G, and H of the row for that device.</li>
 
−
                                    <li>⑨  Read your plates, taking care to use the exact same settings used for your fluorescein measurement.</li>
 
  
  
−
                                </ul>
 
−
                            </li>
 
−
                        <h5><strong>The initial OD600 measurement of our overnight cultures.</strong> </h5>
 
−
                        <img title="demo1" src="https://static.igem.org/mediawiki/2017/1/1a/2017_HUST_China_interlab_img_table1.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                     <div class="col-xs-12">
−
                         <h5><strong>What type of media did you use for this step? </strong> </h5>
+
                         <h4><strong><I>c. Intelligent Simulation</I></strong></h4>
−
                        <p>Luria Bertani </p>
+
                         <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>
−
                         <h5><strong>What type of vessel or container did you use to grow your cells? </strong> </h5>
+
                         <p>The video and figure is here.</p>
−
                         <p>50 ml Falcon tube</p>
+
                         <section class="col-xs-6 col-xs-offset-2">
−
                         <h5><strong>What temperature setting did you use during the measurement?</strong> </h5>
+
                            <figure>
−
                        <p>22℃</p>
+
                                <video controls width="400" height="300" poster="https://static.igem.org/mediawiki/2017/d/db/2017_HUST_China_vedio_poster.png" preload="metadata" aria-describedby="full-descript">
−
                        <h5><strong>What type of 96-well plates did you use?</strong> </h5>
+
                                    <source type="video/webm" src="https://static.igem.org/mediawiki/2017/a/ab/2017_HUST_China_model_vedio4.2.webm" />
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                        <p></p>
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                                    <source type="video/mp4" src="https://static.igem.org/mediawiki/2017/a/af/HUST-modeling4.mp4" />
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                        <h5><strong>Black plates with transparent/clear bottom (preferred)</strong> </h5>
+
                                    <track src="subs/TOS-arabic.srt" kind="subtitles" srclang="ar" label="Arabic" />
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                        <p>Flat</p>
+
                                    <track src="subs/TOS-japanese.srt" kind="subtitles" srclang="jp" label="Japanese" />
−
                        <h5><strong>Measurement </strong> </h5>
+
                                    <track src="subs/TOS-english.srt" kind="subtitles" srclang="en" label="English" />
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                        <ul class="yuandian">
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                                    <track src="subs/TOS-turkish.srt" kind="subtitles" srclang="tr" label="Turkish" />
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                                <li>Measure OD and fluorescence of all samples</li>
+
                                    <track src="subs/TOS-ukrainian.srt" kind="subtitles" srclang="uk" label="Ukrainian" />
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                                <li>Import data into blue cells in Excel (cell measurement) sheets provided</li>
+
                                </video>
−
                        </ul>
+
                            </figure>
−
                        <h5><strong>Suggested Plate Layout for 96-well Plate</strong> </h5>
+
                        </section>
−
                        <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/c/c7/2017_HUST_China_model_ano.png" rel="lightbox-demo" title="my caption"><br />
−
                         <a href="https://static.igem.org/mediawiki/2017/8/87/2017_HUST_China_interlab_img_003.png" rel="lightbox-demo" title="my caption"><br />
+
                             <img src="https://static.igem.org/mediawiki/2017/c/c7/2017_HUST_China_model_ano.png" alt="figure" class="col-xs-8 col-xs-offset-1">
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                             <img title="demo1" src="https://static.igem.org/mediawiki/2017/8/87/2017_HUST_China_interlab_img_003.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
+
 
                         </a>
 
                         </a>
 +
                    </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 20 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 recycle. When it passes 100 units of time, other lanthanide ion release at another position in bottom. Reebot successfully enrich them too. Unfortunately, Reebot also meet waste problem in this situation. So we come to an idea that we can change the recombined protien, to let Reebot catch REE faster. So we repeat LBT sequence. And during modeling analysis, we find the three time LBT is the best.</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 enrich it, then Reebot combine with Si-board. </p>
 
                     </div>
 
                     </div>
 
                 </div>
 
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                    <h3 style="background-color:#ce6b9a;color:#ffffff; padding:10px;letter-spacing:1px;margin-top:0;">Interlab Resultn</h3>
 
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                    <h5 class="col-xs-12"><strong>OD600 Reference Point</strong> </h5>
 
−
                    <img title="demo1" src="https://static.igem.org/mediawiki/2017/5/52/2017_HUST_China_interlab_img_table2.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                    <h5 class="col-xs-12"><strong>Fluorescence standard curve</strong> </h5>
 
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                    <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/>
 
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                        <a href="https://static.igem.org/mediawiki/2017/a/a1/2017_HUST_China_interlab_img_table3.png" rel="lightbox-demo" title="my caption"><br />
 
−
                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/a/a1/2017_HUST_China_interlab_img_table3.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
−
                        </a>
 
−
                    <p class="col-xs-12"><strong>NOTE: 50uM Sample exceeds the range of measurements</strong> </p>
 
−
                    <h5 class="col-xs-12"><strong>OD600 Reference Point</strong> </h5>
 
−
                    <img title="demo1" src="https://static.igem.org/mediawiki/2017/d/d1/2017_HUST_China_interlab_img_table4.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                    <img title="demo1" src="https://static.igem.org/mediawiki/2017/d/df/2017_HUST_China_interlab_img_table5.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                    <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/>
 
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                        <a href="https://static.igem.org/mediawiki/2017/b/b3/2017_HUST_China_interlab_img_table6.png" rel="lightbox-demo" title="my caption"><br />
 
−
                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/b/b3/2017_HUST_China_interlab_img_table6.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
−
                        </a>
 
−
                    <p class="col-xs-12"><strong>Final scaling level determined from medium-high points likely to be less impacted by saturation or pipetting error.</strong> </p>
 
−
                      <p class="col-xs-12"><strong>If needed, you can shift which points are used, but it is likely better to correct instrument settings and protocol.</strong> </p>
 
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                        <a href="https://static.igem.org/mediawiki/2017/b/b8/2017_HUST_China_interlab_img_table7.png" rel="lightbox-demo" title="my caption"><br />
 
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                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/b/b8/2017_HUST_China_interlab_img_table7.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                        </a>
 
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                        <a href="https://static.igem.org/mediawiki/2017/e/ea/2017_HUST_China_interlab_img_table8.png" title="my caption"><br />
 
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                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/e/ea/2017_HUST_China_interlab_img_table8.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                        </a>
 
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                    <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/>
 
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                        <a href="https://static.igem.org/mediawiki/2017/d/d2/2017_HUST_China_interlab_img_table9.png" rel="lightbox-demo" title="my caption"><br />
 
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                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/d/d2/2017_HUST_China_interlab_img_table9.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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                        </a>
 
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                        <a href="https://static.igem.org/mediawiki/2017/2/2b/2017_HUST_China_interlab_img_table10.png" rel="lightbox-demo" title="my caption" style="padding: 10px;"><br />
 
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                            <img title="demo1" src="https://static.igem.org/mediawiki/2017/2/2b/2017_HUST_China_interlab_img_table10.png" alt="demo1" class="col-xs-8 col-xs-offset-2">
 
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             <h3><strong>Acknowledgments:</strong></h3>
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Latest revision as of 03:52, 16 December 2017

Model

「Model」

Overview

We build an ecosystem level model, in order to see how the Reebot diffuse, sense and enrich the lanthanide ions, 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 for 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 Recycled Bacteria
f5 concentration of recycled sensed bacteria
f6 concentration of enriched recycled 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 enriched recycle 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 enriched
LBlever The amount of lanthanide ion enriched 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:

Recycle

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 recycle 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:

Sense and Enrichment

This is the biggest part of our model. We should take lanthanide ion, sensing and enrichment 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 recycled）as f3 and concentration of recycled bacteria (have enriched enough lanthanide ion) as f4.

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

We should also modify the enrichment function. It will only use to sensed bacteria and enriched 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.



figure

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.



figure

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

Result

a. High Concentration Simulation

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.And we will set the Si-board later. 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 enrich the lanthanide ion in the environment. As time goes by, Reebot have enriched more and more lanthanide. At the beginning, the concentration of lanthanide ion is approximately 0.03 unit，and now, it is about 0.003.

When t = 50, we set a 6*8 Si-board on the top. The sensed bacteria and enriched bacteria are recycle to the Si-board. You can see it clearly that the edge of Si-board enriched 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. So, in experiment we change the way Si-Board set, like this way.


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But it's quite difficult to simulate this model, so we still use the old container model
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.



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From the figure, we can see lot of things. In the first 50 units of time, most Reebot quickly enriched enough lanthanide ions, 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 enrich at about a same speed. Then enrichm 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.

b. Low Concentration Simulation

According to the paper, the concentration of lanthanide ion is much lower than expected. So in this simulation, we set concentration of lanthanide ion as 0.001 unit. Other parameters are same as before. Here is the result:


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In this situation, Reebot enriched most of lanthanide ion. But actually, some Reebot are wasted. these Reebot sensed lanthanide ion and expressed LBT and Sitag on their surface. But sensed bacteria is so much that only part of them can enrich enough lanthanide ion. Others only enriched a little and be recycled to Si-board.

c. Intelligent Simulation

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.


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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 20 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 recycle. When it passes 100 units of time, other lanthanide ion release at another position in bottom. Reebot successfully enrich them too. Unfortunately, Reebot also meet waste problem in this situation. So we come to an idea that we can change the recombined protien, to let Reebot catch REE faster. So we repeat LBT sequence. And during modeling analysis, we find the three time LBT is the best.

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 enrich it, then Reebot combine with Si-board.

Acknowledgments: