Team:NYMU-Taipei/ymiq4.html

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       <a href="https://2012.igem.org/Team:NYMU-Taipei"><img src="https://static.igem.org/mediawiki/2012/7/7d/Ymi_header.jpg" border="0"></a>
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       <a href="https://2012.igem.org/Team:NYMU-Taipei"><img src="https://static.igem.org/mediawiki/2012/1/15/Ymi_header1.jpg" border="0"></a>
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<div class="title">Measurements</div>
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<div class="title">Results &amp; References</div>
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   <p><span class="subtitle">The effect of sodium sulfide on Synechococcus SP. PCC 7942 growth rate</span></p>
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   <p><span class="subtitle">Sulfide concentration and the growth of sqr expressing strain Synechococcus sp. PCC 7942</span></p>
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   <p>After thoroughly examined the ability of sqr in Synechococcus SP. PCC 7002, we planned to conduct a series of similar experiments on Synechococcus SP. PCC 7942. Except for the cultivation medium, other growing conditions remained the same. Instinctively, the strain expressing sqr should grow better than the wile type strain.<br />
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    <p align="center"><img src="https://static.igem.org/mediawiki/2012/9/96/Ymiq5-1.png" alt="" width="557" height="299" /><br />
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    </p>
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    <p align="center"><img src="https://static.igem.org/mediawiki/2012/1/16/Ymiq5-2.png" alt="" width="557" height="312" /><br />
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    </p>
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  <p>The  horizontal axis stands for day passed, while the vertical axis is the  absorbance of an OD 730nm. According to the graph, sqr expressing strain grew  much better than wild type strain when sulfide was added into the medium. This  result suggests that sqr is not only expressed but functional in the  cyanobacteria.<br />
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  <p><span class="subtitle">Standard curve</span></p>
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     <p><span class="subtitle">DCMU concentration and cell growth</span></p>
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    <p>This experiment is similar to the second one of Synechococcus SP. PCC 7002 testing series. The main idea was to find the suitable DCMU concentration for Synechococcus SP. PCC 7942. As a matter of fact, both wild type and sqr expressing strain are used in the experiment. <br />
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  <p>We  established a H2S standard curve to quantify H2S concentration inside our experiments.<br />
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  </p></div>
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      <p><span class="subtitle">Sulfide concentration and the growth of sqr expressing strain Synechococcus SP. PCC 7942</span></p>
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  <p><span class="subtitle"> Lab Incubation with Escherichia coli</span></p>
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     <p>It was expected that  SQR expressing strain and wild type counterpart would have different growth  rate under the presence of sulfide compounds. Though sulfide is naturally toxic  to <em>Synechococcus SP. PCC 7942</em>, the  strain with sqr should be able to metabolize sulfide and therefore prosper. As  the result, we analyze H2S amount to detect whether sqr gene work or not. Therefore, we perform Chemical microvolume  turbidimetry method to detect H2S concentration (see <strong>Sulfur Oxide Terminator part</strong>)<br />
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  (1) Different Concentration under 24HR
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     <p align="center"><img src="https://static.igem.org/mediawiki/2012/f/f3/Ymiq7.png" width="510"  /><br />
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    </p>
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  <p>We used different H2S concentration challenge ptrc-sqr transformed E.coli and tested H2S consumption in 24hrs. The result shows that ptrc-sqr transformed E.coli consumed much more H2S compred to the blank control, ptrc-str transformed E.coli. In other words, we believe our sqr gene works!!<br />
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    <br />
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    <br />
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    (2)
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    Different  Concentration under 48HR
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        </p><div class=out style='text-align:center'>
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                  <p align="center"><img src="https://static.igem.org/mediawiki/2012/a/ac/Ymiq8-1.png" width="557" height="362"  /><br />
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      <p><span class="subtitle"> Sulfide oxidation in Escherichia coli expressing sulfide-quinone reductase gene</span></p>
 
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Repots have it that Escherichia coli can express functional sulfide-quinone reductase (SQR). Therefore, we slightly adjusted the previous experiment and applied to the SQR gene from Synechococcus SP. PCC 7002. With methylene blue method, we would test the efficiency of SQR sulfide oxidation. Since such method involved in measurement of optical density, it is more appropriate to perform such experiment on colorless bacteria instead of engineered cyanobacteria strain.
 
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We followed the experiments in 48hrs and analyzed the H2S consumption in different groups. As the same as our expected, in 48hrs, our ptrc-sqr transformed E.coli depleted much more H2S than 24hrs timepoint.<br />
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<br />
<br />
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  </div>
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  <p><span class="subtitle"> Differences Between 24 HR and 48 HR under Same Concentration of H2S</span></p>
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<div align="left">
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  (1) 500mM H2S
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    <p align="center"><img src="https://static.igem.org/mediawiki/2012/0/01/Ymiq9.png" width="628" height="153" /><br />
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    </p>
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  <div class=out style='text-align:center'>
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    <p align="center"><img src="https://static.igem.org/mediawiki/2012/f/fa/Ymiq9-1.png" width="557" height="299" /><br />
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    </p>
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  <p>If  we analyze two group in the same H2S concentration, it&rsquo;s easier to find out  that our ptrc-sqr transformed E.coli consumed H2S dramatically.<br />
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    <br />
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<br />
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    (2)
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250mM H2S  </p>
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  <div class=out style='text-align:center'>
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                  <p align="center"><img src="https://static.igem.org/mediawiki/2012/4/49/Ymiq10.png" width="628" height="153" /><br />
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    </p>
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The  experiment result are the same as previous group, ptrc-sqr transformed E.coli  can consume much more H2S. <br />
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<br />
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<br />
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    (2)
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125mM H2S  </p>
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                  <p align="center"><img src="https://static.igem.org/mediawiki/2012/a/a6/Ymiq11.png" width="628" height="177" /><br />
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    </p>
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  <p>This  diagram illustrates that even at a concentration of 125mM of sodium sulfide,  the difference of consumption rate is obvious between sqr expressing strain and  wild type one.</p>
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  <br />
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  <p><span class="subtitle"> References</span></p>
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<div align="left"><em>Facultative  Anoxygenic Photosynthesis in the Cyanobacterium Oscillatoria limnetica </em>YEHUDA COHEN,* ETANA  PADAN, AND MOSHE SHILO Department of Microbiological Chemistry, The Hebrew  University-Hadassah Medical School, Jerusalem, Israel Received for publication  9 May 1975<br />
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  <br />
 +
  <em>Sulfide  oxidation in gram-negative bacteria by expression of the sulfide–quinone  reductase gene of Rhodobacter capsulatus and by electron transport to  ubiquinone</em> Hiroomi Shibata and Shigeki Kobayashi 2001<br />
 +
  <br />
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  <em>Sulfur metabolism in  Thiorhodoceae. I. Quantitative measurements on growing cells of Chromatium  okenii</em>.  Antonie Leeuwenhoek, 30: 225–238 Trüper, H.G., and Schlegel, H.G. 1964.<br />
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                 <li><a title="Abstract" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq1.html">Abstract</a></li>
                 <li><a title="Abstract" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq1.html">Abstract</a></li>
                 <li><a title="Methods" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq2.html">Methods</a></li>
                 <li><a title="Methods" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq2.html">Methods</a></li>
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                 <li><a title="Measurements" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq3.html">Measurements</a></li>
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                 <li><a title="Experiments" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq3.html">Experiments</a></li>
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                 <li><a title="Results & References" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq4.html">Results &amp; References</a></li>
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                 <li><a title="Results & References" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq4.html">Results &amp; References</a></li><li><a title="Further Experiments after Asia Jamboree" href="https://2012.igem.org/Team:NYMU-Taipei/ymiq5.html">Further Experiments after Asia Jamboree</a></li>
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                 <li><a title="Methods & Materials" href="https://2012.igem.org/Team:NYMU-Taipei/ymic3.html">Methods & Materials</a></li>
                 <li><a title="Methods & Materials" href="https://2012.igem.org/Team:NYMU-Taipei/ymic3.html">Methods & Materials</a></li>
                 <li><a title="Results & Discussion" href="https://2012.igem.org/Team:NYMU-Taipei/ymic4.html">Results & Discussion</a></li>
                 <li><a title="Results & Discussion" href="https://2012.igem.org/Team:NYMU-Taipei/ymic4.html">Results & Discussion</a></li>
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                 <li><a title="Conclusioin & References" href="https://2012.igem.org/Team:NYMU-Taipei/ymic5.html">Conclusioin & References</a></li>
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                 <li><a title="Conclusion & References" href="https://2012.igem.org/Team:NYMU-Taipei/ymic5.html">Conclusion & References</a></li>
                  
                  
             </ul>
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Latest revision as of 01:49, 27 October 2012

NYMU iGEM

Results & References

Sulfide concentration and the growth of sqr expressing strain Synechococcus sp. PCC 7942



The horizontal axis stands for day passed, while the vertical axis is the absorbance of an OD 730nm. According to the graph, sqr expressing strain grew much better than wild type strain when sulfide was added into the medium. This result suggests that sqr is not only expressed but functional in the cyanobacteria.


Standard curve


We established a H2S standard curve to quantify H2S concentration inside our experiments.


Lab Incubation with Escherichia coli

(1) Different Concentration under 24HR


We used different H2S concentration challenge ptrc-sqr transformed E.coli and tested H2S consumption in 24hrs. The result shows that ptrc-sqr transformed E.coli consumed much more H2S compred to the blank control, ptrc-str transformed E.coli. In other words, we believe our sqr gene works!!


(2) Different Concentration under 48HR


We followed the experiments in 48hrs and analyzed the H2S consumption in different groups. As the same as our expected, in 48hrs, our ptrc-sqr transformed E.coli depleted much more H2S than 24hrs timepoint.


Differences Between 24 HR and 48 HR under Same Concentration of H2S

(1) 500mM H2S



If we analyze two group in the same H2S concentration, it’s easier to find out that our ptrc-sqr transformed E.coli consumed H2S dramatically.


(2) 250mM H2S


The experiment result are the same as previous group, ptrc-sqr transformed E.coli can consume much more H2S.


(2) 125mM H2S


This diagram illustrates that even at a concentration of 125mM of sodium sulfide, the difference of consumption rate is obvious between sqr expressing strain and wild type one.


References

Facultative Anoxygenic Photosynthesis in the Cyanobacterium Oscillatoria limnetica YEHUDA COHEN,* ETANA PADAN, AND MOSHE SHILO Department of Microbiological Chemistry, The Hebrew University-Hadassah Medical School, Jerusalem, Israel Received for publication 9 May 1975

Sulfide oxidation in gram-negative bacteria by expression of the sulfide–quinone reductase gene of Rhodobacter capsulatus and by electron transport to ubiquinone Hiroomi Shibata and Shigeki Kobayashi 2001

Sulfur metabolism in Thiorhodoceae. I. Quantitative measurements on growing cells of Chromatium okenii. Antonie Leeuwenhoek, 30: 225–238 Trüper, H.G., and Schlegel, H.G. 1964.