Difference between revisions of "Team:HZAU-China/InterLab"

 
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                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Improve">Improve</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Improve">Improve</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/InterLab">InterLab</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/InterLab">InterLab</a>
 +
                <a class="item" href="https://2018.igem.org/Team:HZAU-China/Notebook">Notebook</a>
 +
 
             </li>
 
             </li>
 
             <li class="hiLight shortName">
 
             <li class="hiLight shortName">
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                     <span class="xjtPic"></span>
 
                     <span class="xjtPic"></span>
 
                 </a>
 
                 </a>
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                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Safety">Safty</a>
+
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Safety">Safety</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Human_Practices">Human Practices</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Human_Practices">Human Practices</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Public_Engagement">Public Engagement</a>
 
                 <a class="item" href="https://2018.igem.org/Team:HZAU-China/Public_Engagement">Public Engagement</a>
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         <!-- 正文 -->
 
         <!-- 正文 -->
 
         <div class="zhengwen">
 
         <div class="zhengwen">
−
             <img class="daimg" src="https://static.igem.org/mediawiki/2018/7/70/T--HZAU-China--hezhao1.png" alt="">
+
             <p style="padding-top:25px;">The HZAU-China team also participated in the InterLab project as we did
 +
                in the past years. At
 +
                first we read and analyzed the protocol provided by the headquarters carefully, then we checked out if
 +
                we had
 +
                all the essential material and equipment. After everything was well prepared, we formally carried
 +
                out the experiment.</p>
 
             <div id="float01" class="cur">
 
             <div id="float01" class="cur">
−
                <div class="h1">biaotiyi</div>
 
  
 +
                <div class="h1">Calibration</div>
 +
                <div class="h2">OD600 Reference Point</div>
 +
                <p>To obtain the OD600 reference point of a particular plate reader is rather necessary since we can
 +
                    only obtain ABS from a plate reader which is different from OD gained from a spectrophotometer. The
 +
                    ABS data obtained from a plate reader is volume-dependent while the sample of a spectrophotometer
 +
                    are volume-fixed.</p>
 +
                <p>As the protocol instructed, we added 100μL LUDOX solution into wells A1, B1, C1, D1 while added
 +
                    100μL ddH<sub>2</sub>O into wells A2, B2, C2, D2. Then, we measured OD600 of the samples in the
 +
                    plate reader
 +
                    (Table 1).</p>
 +
                <p style="width: 100%; text-align: center !important;"><b>Table 1.</b> The result of OD600
 +
                    reference point measuring. </p>
 +
                <div style="width: 60%; margin: 30px auto">
 +
                    <!-- <div class="table-responsive"> -->
 +
                    <table class="table table-bordered table-hover">
 +
                        <thead>
 +
                            <tr>
 +
                                <th></th>
 +
                                <th>LUDOX CL-X</th>
 +
                                <th>H<sub>2</sub>O</th>
 +
                            </tr>
 +
                        </thead>
 +
                        <tbody>
 +
                            <tr>
 +
                                <td>Replicate 1</td>
 +
                                <td class="success">0.057</td>
 +
                                <td class="success">0.036</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 2</td>
 +
                                <td class="success">0.055</td>
 +
                                <td class="success">0.039</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 3</td>
 +
                                <td class="success">0.056</td>
 +
                                <td class="success">0.038</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 4</td>
 +
                                <td class="success">0.056</td>
 +
                                <td class="success">0.039</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Arith. Mean</td>
 +
                                <td class="warning">0.056</td>
 +
                                <td class="warning">0.038</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Corrected Abs600</td>
 +
                                <td class="warning">0.018</td>
 +
                                <td></td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Reference OD600</td>
 +
                                <td class="warning">0.063</td>
 +
                                <td></td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>OD600/Abs600</td>
 +
                                <td class="warning">3.500</td>
 +
                                <td></td>
 +
                            </tr>
 +
                        </tbody>
 +
                    </table>
 +
                    <!-- </div> -->
 +
                </div>
 +
 +
                <p>The data showed that the OD600 reference point of our plate reader is 3.500.</p>
 +
                <div class="h2">Particle Standard Curve</div>
 +
                <p>The particle standard curve experiment aims to obtain the relationship curve between the ABS600 of
 +
                    the solution and the concentration of bacteria. To achieve this goal, we need to find a kind of
 +
                    material which resembles bacteria very much. The headquarters sent us with Silica Beads, a kind of
 +
                    little
 +
                    particle which met our needs.</p>
 +
                <p>We made up a series of solution of different but continuous magnitude. To make the experiment more
 +
                    precise, we also set up another three parallel groups (Figure 1).</p>
 +
                <div style="width: 60%; margin: 30px auto">
 +
                    <img src="https://static.igem.org/mediawiki/2018/3/38/T--HZAU-China--interlab01.png" width=100% alt="">
 +
                    <div style="height:40px; width:100%"></div>
 +
                    <img src="https://static.igem.org/mediawiki/2018/8/87/T--HZAU-China--interlab02.png" width=100% alt="">
 +
                </div>
 +
                <p style="width: 100%; text-align: center !important;"><b>Figure 1.</b> The result of Particle Standard Curve.</p>
 +
                <div class="h2">Fluorescence Standard Curve</div>
 +
 +
                <p>The fluorescence standard curve experiment targets to obtain the relationship curve between the
 +
                    strength of fluorescence and the concentration of fluorescein which has similar excitation and
 +
                    emission properties to GFP. </p>
 +
                <p>We prepared a series of fluorescein solution of different but continuous magnitude. As every
 +
                    experiment required, we also set up 4 parallel groups. Then we measured the strength of the
 +
                    fluorescence of each sample (Figure 2).</p>
 +
                <div style="width: 60%; margin: 30px auto">
 +
                    <img src="https://static.igem.org/mediawiki/2018/d/dd/T--HZAU-China--interlab03.png" width=100% alt="">
 +
                    <div style="height:40px; width:100%"></div>
 +
                    <img src="https://static.igem.org/mediawiki/2018/8/82/T--HZAU-China--interlab04.png" width=100% alt="">
 +
                </div>
 +
                <p style="width: 100%; text-align: center !important;"><b>Figure 2.</b> The result of Fluorescence Standard Curve.</p>
 
             </div>
 
             </div>
 
             <div id="float02">
 
             <div id="float02">
−
                 <div class="h1">biaotier</div>
+
                 <div class="h1">Cell Measurement</div>
 +
                <p>Cell measurement was the most complicated experiment we've ever countered in the whole InterLab. It
 +
                    nearly took us a whole week to complete and diagnose the data. Each step of the experiment was so
 +
                    crucial that it could impact the result to a detectable extent.
 +
                </p>
 +
                <p>
 +
                </p>
 +
                <p>However, we fixed up all the mistakes and completed the experiment satisfactorily (Table 2).</p>
 +
                <p style="width: 100%; text-align: center !important;"><b>Table 2.</b> Our raw data of cell measurement.</p>
 +
                <div style="width: 80%; margin: 30px auto">
 +
                    <img src="https://static.igem.org/mediawiki/2018/8/8e/T--HZAU-China--interlab05.png" width=100% alt="">
 +
                    <div style="height:40px; width:100%"></div>
 +
                    <img src="https://static.igem.org/mediawiki/2018/3/3d/T--HZAU-China--interlab06.png" width=100% alt="">
 +
                </div>
  
 +
                <p>After repeated experiments we chose the best data and upload them to headquarters.</p>
 +
                <div class="h2">Colony Forming Units per 0.1 OD600 E. coli cultures</div>
 +
                <p>The CFU experiment is another challenging mission for its large amount of work and complicated
 +
                    operations.
 +
                </p>
 +
                <p> We diluted the bacteria solution to OD600 = 0.1, then made up a sort of magnitude as the protocol
 +
                    required.
 +
                </p>
 +
                <p> After reconducting the experiment for three times, we successfully achieved the data required by
 +
                    headquarters
 +
                    and finished the InterLab satisfactorily (Table 3).
 +
                </p>
 +
                <p style="width: 100%; text-align: center !important;"><b>Table 3.</b> Our raw data of cell measurement.</p>
 +
                <div style="width: 60%; margin: 30px auto">
 +
                    <!-- <div class="table-responsive"> -->
 +
                    <table class="table table-bordered table-hover">
 +
                        <thead>
 +
                            <tr>
 +
                                <th></th>
 +
                                <th>Replicate 1</th>
 +
                                <th>Replicate 2</th>
 +
                                <th>Replicate 3</th>
 +
                            </tr>
 +
                        </thead>
 +
                        <tbody>
 +
                            <tr>
 +
                                <td>Positive 1</td>
 +
                                <td>0.084</td>
 +
                                <td>0.0945</td>
 +
                                <td>0.0875</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Positive 2</td>
 +
                                <td>0.098</td>
 +
                                <td>0.084</td>
 +
                                <td>0.0875</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Negative 1</td>
 +
                                <td>0.0945</td>
 +
                                <td>0.084</td>
 +
                                <td>0.0875</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Negative 2</td>
 +
                                <td>0.1015</td>
 +
                                <td>0.098</td>
 +
                                <td>0.1015</td>
 +
                            </tr>
 +
                        </tbody>
 +
                    </table>
 +
                    <!-- </div> -->
 +
 +
                    <!-- <div class="table-responsive"> -->
 +
                    <table class="table table-bordered table-hover">
 +
                        <caption class="text-center">Positive 1</caption>
 +
                        <tbody>
 +
                            <tr>
 +
                                <td>Replicate 1</td>
 +
                                <td><sup>a</sup>TMTC</td>
 +
                                <td>11</td>
 +
                                <td>1</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 2</td>
 +
                                <td>206</td>
 +
                                <td>18</td>
 +
                                <td>6</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 3</td>
 +
                                <td>174</td>
 +
                                <td>17</td>
 +
                                <td>2</td>
 +
                            </tr>
 +
                        </tbody>
 +
                    </table>
 +
                    <!-- </div> -->
 +
                    <!-- <div class="table-responsive"> -->
 +
                    <table class="table table-bordered table-hover">
 +
                        <caption class="text-center">Positive 2</caption>
 +
                        <tbody>
 +
                            <tr>
 +
                                <td>Replicate 1</td>
 +
                                <td>TMTC</td>
 +
                                <td>9</td>
 +
                                <td>4</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 2</td>
 +
                                <td>228</td>
 +
                                <td>25</td>
 +
                                <td>8</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 3</td>
 +
                                <td>278</td>
 +
                                <td>2</td>
 +
                                <td>0</td>
 +
                            </tr>
 +
                        </tbody>
 +
                    </table>
 +
                    <!-- </div> -->
 +
                    <!-- <div class="table-responsive"> -->
 +
                    <table class="table table-bordered table-hover">
 +
                        <caption class="text-center">Negative 1</caption>
 +
                        <tbody>
 +
                            <tr>
 +
                                <td>Replicate 1</td>
 +
                                <td>TMTC</td>
 +
                                <td>23</td>
 +
                                <td>1</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 2</td>
 +
                                <td>370</td>
 +
                                <td>28</td>
 +
                                <td>10</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 3</td>
 +
                                <td>TMTC</td>
 +
                                <td>11</td>
 +
                                <td>1</td>
 +
                            </tr>
 +
                        </tbody>
 +
                    </table>
 +
                    <!-- </div> -->
 +
                    <!-- <div class="table-responsive"> -->
 +
                    <table class="table table-bordered table-hover">
 +
                        <caption class="text-center">Negative 2</caption>
 +
                        <tbody>
 +
                            <tr>
 +
                                <td>Replicate 1</td>
 +
                                <td>TMTC</td>
 +
                                <td>89</td>
 +
                                <td>1</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 2</td>
 +
                                <td>TMTC</td>
 +
                                <td>16</td>
 +
                                <td>0</td>
 +
                            </tr>
 +
                            <tr>
 +
                                <td>Replicate 3</td>
 +
                                <td>TMTC</td>
 +
                                <td>48</td>
 +
                                <td>4</td>
 +
                            </tr>
 +
                        </tbody>
 +
                    </table>
 +
                    <!-- </div> -->
 +
                </div>
 +
                <p><sup>a</sup>TMTC in our forms means the colony forming units in this plate is too much to
 +
                    count (over 300).</p>
 
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             <div id="float03">
 
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                 <div class="h1">Summary</div>
−
                  
+
                 <p>With this, we are delighted to announce that we completed the work on punctual with satisfaction
 +
                    from Measurement Committee.
 +
                </p>
 +
                <p style="padding-bottom:25px;"> The only suggestion we want to submit is that, in the Cell Measurement
 +
                    Experiment, we don’t think
 +
                    it’s correct to put the sample at 0 hour on ice. Because even if you put it on ice, the bacteria
 +
                    will also definitely grow, which results in high systematical error in the experiment.
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                </p>
 
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                 <p class="daohanger">Summary</p>
 
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                 <a>
 
                     <span>Back&nbsp;to&nbsp;Top</span>
 
                     <span>Back&nbsp;to&nbsp;Top</span>
 
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Latest revision as of 20:59, 17 October 2018

The HZAU-China team also participated in the InterLab project as we did in the past years. At first we read and analyzed the protocol provided by the headquarters carefully, then we checked out if we had all the essential material and equipment. After everything was well prepared, we formally carried out the experiment.

Calibration
OD600 Reference Point

To obtain the OD600 reference point of a particular plate reader is rather necessary since we can only obtain ABS from a plate reader which is different from OD gained from a spectrophotometer. The ABS data obtained from a plate reader is volume-dependent while the sample of a spectrophotometer are volume-fixed.

As the protocol instructed, we added 100μL LUDOX solution into wells A1, B1, C1, D1 while added 100μL ddH2O into wells A2, B2, C2, D2. Then, we measured OD600 of the samples in the plate reader (Table 1).

Table 1. The result of OD600 reference point measuring.

LUDOX CL-X H2O
Replicate 1 0.057 0.036
Replicate 2 0.055 0.039
Replicate 3 0.056 0.038
Replicate 4 0.056 0.039
Arith. Mean 0.056 0.038
Corrected Abs600 0.018
Reference OD600 0.063
OD600/Abs600 3.500

The data showed that the OD600 reference point of our plate reader is 3.500.

Particle Standard Curve

The particle standard curve experiment aims to obtain the relationship curve between the ABS600 of the solution and the concentration of bacteria. To achieve this goal, we need to find a kind of material which resembles bacteria very much. The headquarters sent us with Silica Beads, a kind of little particle which met our needs.

We made up a series of solution of different but continuous magnitude. To make the experiment more precise, we also set up another three parallel groups (Figure 1).

Figure 1. The result of Particle Standard Curve.

Fluorescence Standard Curve

The fluorescence standard curve experiment targets to obtain the relationship curve between the strength of fluorescence and the concentration of fluorescein which has similar excitation and emission properties to GFP.

We prepared a series of fluorescein solution of different but continuous magnitude. As every experiment required, we also set up 4 parallel groups. Then we measured the strength of the fluorescence of each sample (Figure 2).

Figure 2. The result of Fluorescence Standard Curve.

Cell Measurement

Cell measurement was the most complicated experiment we've ever countered in the whole InterLab. It nearly took us a whole week to complete and diagnose the data. Each step of the experiment was so crucial that it could impact the result to a detectable extent.

However, we fixed up all the mistakes and completed the experiment satisfactorily (Table 2).

Table 2. Our raw data of cell measurement.

After repeated experiments we chose the best data and upload them to headquarters.

Colony Forming Units per 0.1 OD600 E. coli cultures

The CFU experiment is another challenging mission for its large amount of work and complicated operations.

We diluted the bacteria solution to OD600 = 0.1, then made up a sort of magnitude as the protocol required.

After reconducting the experiment for three times, we successfully achieved the data required by headquarters and finished the InterLab satisfactorily (Table 3).

Table 3. Our raw data of cell measurement.

Replicate 1 Replicate 2 Replicate 3
Positive 1 0.084 0.0945 0.0875
Positive 2 0.098 0.084 0.0875
Negative 1 0.0945 0.084 0.0875
Negative 2 0.1015 0.098 0.1015
Positive 1
Replicate 1 aTMTC 11 1
Replicate 2 206 18 6
Replicate 3 174 17 2
Positive 2
Replicate 1 TMTC 9 4
Replicate 2 228 25 8
Replicate 3 278 2 0
Negative 1
Replicate 1 TMTC 23 1
Replicate 2 370 28 10
Replicate 3 TMTC 11 1
Negative 2
Replicate 1 TMTC 89 1
Replicate 2 TMTC 16 0
Replicate 3 TMTC 48 4

aTMTC in our forms means the colony forming units in this plate is too much to count (over 300).

Summary

With this, we are delighted to announce that we completed the work on punctual with satisfaction from Measurement Committee.

The only suggestion we want to submit is that, in the Cell Measurement Experiment, we don’t think it’s correct to put the sample at 0 hour on ice. Because even if you put it on ice, the bacteria will also definitely grow, which results in high systematical error in the experiment.

Experiments Improve
InterLab

Calibration

Cell Measurement

Summary

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Notebook