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<p style="padding-top:25px;">The HZAU-China team also participated in the InterLab project as we did | <p style="padding-top:25px;">The HZAU-China team also participated in the InterLab project as we did | ||
in the past years. At | in the past years. At | ||
− | first we read and analyzed the protocol provided by the headquarters carefully, then we checked out if we had | + | 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 | all the essential material and equipment. After everything was well prepared, we formally carried | ||
out the experiment.</p> | out the experiment.</p> | ||
Line 714: | Line 715: | ||
are volume-fixed.</p> | are volume-fixed.</p> | ||
<p>As the protocol instructed, we added 100μL LUDOX solution into wells A1, B1, C1, D1 while added | <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 | + | 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> | (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 style="width: 60%; margin: 30px auto"> | ||
<!-- <div class="table-responsive"> --> | <!-- <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> --> | ||
</div> | </div> | ||
− | + | ||
<p>The data showed that the OD600 reference point of our plate reader is 3.500.</p> | <p>The data showed that the OD600 reference point of our plate reader is 3.500.</p> | ||
<div class="h2">Particle Standard Curve</div> | <div class="h2">Particle Standard Curve</div> | ||
<p>The particle standard curve experiment aims to obtain the relationship curve between the ABS600 of | <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 | 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 | + | material which resembles bacteria very much. The headquarters sent us with Silica Beads, a kind of |
+ | little | ||
particle which met our needs.</p> | particle which met our needs.</p> | ||
<p>We made up a series of solution of different but continuous magnitude. To make the experiment more | <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> | + | precise, we also set up another three parallel groups (Figure 1).</p> |
<div style="width: 60%; margin: 30px auto"> | <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=""> | <img src="https://static.igem.org/mediawiki/2018/3/38/T--HZAU-China--interlab01.png" width=100% alt=""> | ||
Line 785: | Line 790: | ||
<img src="https://static.igem.org/mediawiki/2018/8/87/T--HZAU-China--interlab02.png" width=100% alt=""> | <img src="https://static.igem.org/mediawiki/2018/8/87/T--HZAU-China--interlab02.png" width=100% alt=""> | ||
</div> | </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> | <div class="h2">Fluorescence Standard Curve</div> | ||
− | + | ||
<p>The fluorescence standard curve experiment targets to obtain the relationship curve between the | <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 | strength of fluorescence and the concentration of fluorescein which has similar excitation and | ||
Line 793: | Line 798: | ||
<p>We prepared a series of fluorescein solution of different but continuous magnitude. As every | <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 | experiment required, we also set up 4 parallel groups. Then we measured the strength of the | ||
− | fluorescence of each sample(Figure 2).</p> | + | fluorescence of each sample (Figure 2).</p> |
<div style="width: 60%; margin: 30px auto"> | <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=""> | <img src="https://static.igem.org/mediawiki/2018/d/dd/T--HZAU-China--interlab03.png" width=100% alt=""> | ||
Line 799: | Line 804: | ||
<img src="https://static.igem.org/mediawiki/2018/8/82/T--HZAU-China--interlab04.png" width=100% alt=""> | <img src="https://static.igem.org/mediawiki/2018/8/82/T--HZAU-China--interlab04.png" width=100% alt=""> | ||
</div> | </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"> | ||
Line 806: | Line 812: | ||
crucial that it could impact the result to a detectable extent. | crucial that it could impact the result to a detectable extent. | ||
</p> | </p> | ||
− | <p> | + | <p> |
</p> | </p> | ||
− | <p>However, we fixed up all the mistakes and completed the experiment satisfactorily(Table 2).</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"> | <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=""> | <img src="https://static.igem.org/mediawiki/2018/8/8e/T--HZAU-China--interlab05.png" width=100% alt=""> | ||
Line 814: | Line 821: | ||
<img src="https://static.igem.org/mediawiki/2018/3/3d/T--HZAU-China--interlab06.png" width=100% alt=""> | <img src="https://static.igem.org/mediawiki/2018/3/3d/T--HZAU-China--interlab06.png" width=100% alt=""> | ||
</div> | </div> | ||
+ | |||
<p>After repeated experiments we chose the best data and upload them to headquarters.</p> | <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> | <div class="h2">Colony Forming Units per 0.1 OD600 E. coli cultures</div> | ||
Line 820: | Line 828: | ||
</p> | </p> | ||
<p> We diluted the bacteria solution to OD600 = 0.1, then made up a sort of magnitude as the protocol | <p> We diluted the bacteria solution to OD600 = 0.1, then made up a sort of magnitude as the protocol | ||
− | required. | + | required. |
</p> | </p> | ||
− | <p> After reconducting the experiment for three times, we successfully achieved the data required by headquarters | + | <p> After reconducting the experiment for three times, we successfully achieved the data required by |
− | and finished the InterLab satisfactorily(Table 3). | + | headquarters |
+ | and finished the InterLab satisfactorily (Table 3). | ||
</p> | </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 style="width: 60%; margin: 30px auto"> | ||
<!-- <div class="table-responsive"> --> | <!-- <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> --> | ||
<!-- <div class="table-responsive"> --> | <!-- <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> --> | ||
<!-- <div class="table-responsive"> --> | <!-- <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> --> | ||
<!-- <div class="table-responsive"> --> | <!-- <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> --> | ||
<!-- <div class="table-responsive"> --> | <!-- <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> --> | ||
</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> | ||
</div> | </div> | ||
<div id="float03"> | <div id="float03"> | ||
<div class="h1">Summary</div> | <div class="h1">Summary</div> | ||
− | <p> | + | <p>With this, we are delighted to announce that we completed the work on punctual with satisfaction |
− | + | from Measurement Committee. | |
</p> | </p> | ||
<p style="padding-bottom:25px;"> The only suggestion we want to submit is that, in the Cell Measurement | <p style="padding-bottom:25px;"> The only suggestion we want to submit is that, in the Cell Measurement |
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.
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.
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.
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 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.
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 |
Replicate 1 | aTMTC | 11 | 1 |
Replicate 2 | 206 | 18 | 6 |
Replicate 3 | 174 | 17 | 2 |
Replicate 1 | TMTC | 9 | 4 |
Replicate 2 | 228 | 25 | 8 |
Replicate 3 | 278 | 2 | 0 |
Replicate 1 | TMTC | 23 | 1 |
Replicate 2 | 370 | 28 | 10 |
Replicate 3 | TMTC | 11 | 1 |
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).
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.