Difference between revisions of "Team:Jilin China/InterLab"

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<ul style="line-height: 26px;">
 
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<li>Positive Control  (BBa_I20270)</li>
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<li>Positive Control  (BBa_I20270): well 20B</li>
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<li>Negative Control  (BBa_R0040)</li>
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<li>Negative Control  (BBa_R0040): well 20D</li>
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<li>Test Device 1    (BBa_J364000)</li>
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<li>Test Device 1    (BBa_J364000): well 20F</li>
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<li>Test Device 2    (BBa_J364001)</li>
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<li>Test Device 2    (BBa_J364001): well 20H</li>
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<li>Test Device 3    (BBa_J364002)</li>
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<li>Test Device 3    (BBa_J364002): well 20J</li>
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<li>Test Device 4    (BBa_J364003)</li>
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<li>Test Device 4    (BBa_J364003): well 20L</li>
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<li>Test Device 5    (BBa_J364004)</li>
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<li>Test Device 5    (BBa_J364004): well 20N</li>
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<li>Test Device 6    (BBa_J364005)</li>
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<li>Test Device 6    (BBa_J364005): well 20P</li>
 
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<div class="h1_title">Result</div>
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<div class="h1_title">The raw data</div>
 
<div class="h1_content">
 
<div class="h1_content">
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<p>We have sent our data sheets to measurement AT igem DOT org before 29th September.</p>
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<p>The sheets are as followed:</p>
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<div class="pic_box">
 
<div class="pic_box">
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<strong>OD600 reference point</strong><br />
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<strong>1.OD600 reference point</strong><br />
 
<img src="https://static.igem.org/mediawiki/2017/8/85/T--Jilin_China--interlab1.png" />
 
<img src="https://static.igem.org/mediawiki/2017/8/85/T--Jilin_China--interlab1.png" />
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</div>
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<br /><br />
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<div class="pic_box">
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<strong>Fluorescein standard curve</strong><br />
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<strong>2.Fluorescein standard curve</strong><br />
 
<img src="https://static.igem.org/mediawiki/2017/4/4f/T--Jilin_China--interlab2.png" /><br /><br />
 
<img src="https://static.igem.org/mediawiki/2017/4/4f/T--Jilin_China--interlab2.png" /><br /><br />
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<img src="https://static.igem.org/mediawiki/2017/f/f7/T--Jilin_China--interlab3.png" /><br /><br />
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<img src="https://static.igem.org/mediawiki/2017/f/f7/T--Jilin_China--interlab3.png" width="40%" />
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<img src="https://static.igem.org/mediawiki/2017/thumb/9/9a/T--Jilin_China--interlab_log_scale_pic.png/800px-T--Jilin_China--interlab_log_scale_pic.png" /><br /><br />
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<img src="https://static.igem.org/mediawiki/2017/9/96/T--Jilin_China--interlab4.png" width="40%" />
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</div>
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<br /><br />
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<div class="pic_box">
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<strong>Raw Plate Reader Measurements</strong><br />
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<strong>3.Raw Plate Reader Measurements</strong><br />
 
Dilution Calculation:<br />
 
Dilution Calculation:<br />
 
<img src="https://static.igem.org/mediawiki/2017/2/26/T--Jilin_China--interlab5.jpg" /><br /><br />
 
<img src="https://static.igem.org/mediawiki/2017/2/26/T--Jilin_China--interlab5.jpg" /><br /><br />
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<img src="https://static.igem.org/mediawiki/2017/b/b4/T--Jilin_China--interlab6.jpg" /><br /><br />
 
<img src="https://static.igem.org/mediawiki/2017/b/b4/T--Jilin_China--interlab6.jpg" /><br /><br />
 
<img src="https://static.igem.org/mediawiki/2017/e/e8/T--Jilin_China--interlab7.jpg" />
 
<img src="https://static.igem.org/mediawiki/2017/e/e8/T--Jilin_China--interlab7.jpg" />
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<img src="https://static.igem.org/mediawiki/2017/e/e0/T--Jilin_China--interlab9.jpg" /><br /><br />
 
<img src="https://static.igem.org/mediawiki/2017/e/e0/T--Jilin_China--interlab9.jpg" /><br /><br />
 
</div>
 
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</div>
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<div class="h1_title">Data analysis</div>
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<div class="h1_content">
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<strong>1.Abs600</strong>
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<div class="pic_box">
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<img src="https://static.igem.org/mediawiki/2017/0/07/T--Jilin_China--interlab10.png" /><br />
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fig.1a<br /><br />
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<img src="https://static.igem.org/mediawiki/2017/8/87/T--Jilin_China--interlab11.png" /><br />
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fig.1b<br /><br />
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</div>
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<p>fig.1a&b The absorbance at 600nm over time. Samples were collected every two hours and measured Abs600 by using the plate reader.</p>
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<strong>2.Fluorescence</strong>
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<div class="pic_box">
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<img src="https://static.igem.org/mediawiki/2017/9/99/T--Jilin_China--interlab12.png" /><br />
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fig.2a<br /><br />
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<img src="https://static.igem.org/mediawiki/2017/2/21/T--Jilin_China--interlab13.png" /><br />
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fig.2b<br /><br />
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</div>
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<p>fig.2a&b The change of fluorescence over time. Samples were collected every two hours and measured their fluorescence by using the plate reader.</p>
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<strong>3.Contrast between different promoters</strong>
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<div class="pic_box">
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<img src="https://static.igem.org/mediawiki/2017/9/99/T--Jilin_China--interlab14.png" /><br />
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fig.3 The value of Fluorescein/OD600 of every test device in different time.
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Revision as of 10:31, 31 October 2017

Introduction

Reliable and repeatable measurement is a key component to all engineering disciplines. The same holds true for synthetic biology, which has also been called engineering biology. However, the ability to repeat measurements in different labs has been difficult. The Measurement Committee, through the InterLab study, has been developing a robust measurement procedure for green fluorescent protein (GFP) over the last three years. We chose GFP as the measurement marker for this study since it's one of the most used markers in synthetic biology and, as a result, most laboratories are equipped to measure this protein. We think it is a great chance for our team to make our own contribution to this project.

Device we received

  • Positive Control (BBa_I20270): well 20B
  • Negative Control (BBa_R0040): well 20D
  • Test Device 1 (BBa_J364000): well 20F
  • Test Device 2 (BBa_J364001): well 20H
  • Test Device 3 (BBa_J364002): well 20J
  • Test Device 4 (BBa_J364003): well 20L
  • Test Device 5 (BBa_J364004): well 20N
  • Test Device 6 (BBa_J364005): well 20P

Protocol

We strictly followed the protocol provided by iGEM. We measured the experiment of OD600 reference point and fitted the fluorescence standard curve first and then detected GFP expression in our E.coli with the plate reader Synergy HT from BioTek.

The raw data

We have sent our data sheets to measurement AT igem DOT org before 29th September.

The sheets are as followed:

1.OD600 reference point


2.Fluorescein standard curve




3.Raw Plate Reader Measurements
Dilution Calculation:


Fluorescence Raw Readings:



Abs600 Raw Readings




Data analysis
1.Abs600

fig.1a


fig.1b

fig.1a&b The absorbance at 600nm over time. Samples were collected every two hours and measured Abs600 by using the plate reader.

2.Fluorescence

fig.2a


fig.2b

fig.2a&b The change of fluorescence over time. Samples were collected every two hours and measured their fluorescence by using the plate reader.

3.Contrast between different promoters

fig.3 The value of Fluorescein/OD600 of every test device in different time.

Conclusion

As our results showed, promoter J23101 was the strongest among the 3 promoters, which showed the highest GFP fluorescence. Promoter J23106 was moderate, and promoter J23117 was the lowest.

Acknowledgement

We really appreciated Prof. Yubin Ge for providing us the permission to use the plate reader from his lab. Besides, PhD candidate Cheng Hu taught us how to use a plate reader. BioTek service engineer Jeremy Gagne told us some settings of the Synergy HT, which we couldn't find from the setting panel.