Difference between revisions of "Team:Toronto/Results"
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| − | <div class="content" id="content-main"><div class="row"><div class="col col-lg-8 col-md-12"><div class="content-main"> | + | <div class="content" id="content-main"><div class="row"><div class="col col-lg-8 col-md-12"><div class="content-main"><h1 id="results">Results</h1> |
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| − | <h1 id="results">Results</h1> | + | |
<h3 id="success">Success</h3> | <h3 id="success">Success</h3> | ||
<ul> | <ul> | ||
<li>Both <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048002">Long GolS P118A (BBa_K2048002)</a> and <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048001">Long GolS (BBa_K2048001)</a> constructs responded to gold concentrations in a dose-responsive manner up to 1uM </li> | <li>Both <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048002">Long GolS P118A (BBa_K2048002)</a> and <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048001">Long GolS (BBa_K2048001)</a> constructs responded to gold concentrations in a dose-responsive manner up to 1uM </li> | ||
| − | <li><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048002">Long GolS P118A (BBa_K2048002)</a> is more responsive to gold than copper | + | <li><p><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048002">Long GolS P118A (BBa_K2048002)</a> is more responsive to gold than copper |
| − | <img src="https://static.igem.org/mediawiki/parts/6/60/Igem_2016_Gold_Copper_Cell_Assay.jpeg" alt="alt text"></li> | + | <img src="https://static.igem.org/mediawiki/parts/6/60/Igem_2016_Gold_Copper_Cell_Assay.jpeg" alt="alt text"></p> |
| − | <li><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048001">Long GolS (BBa_K2048001)</a> is more responsive to both gold and copper concentrations than York 2013 construct (BBa_K1127008)</li> | + | <center><em>Figure 1: CPRG assay results after 12 hours of incubation</em></center> |
| + | </li> | ||
| + | <li><p><a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048001">Long GolS (BBa_K2048001)</a> is more responsive to both gold and copper concentrations than York 2013 construct (BBa_K1127008)</p> | ||
| + | </li> | ||
</ul> | </ul> | ||
| + | <h3 id="discussion">Discussion</h3> | ||
| + | <p><img src="https://static.igem.org/mediawiki/parts/5/56/2016_iGem_Toronto_GolS_3D.png" alt="alt text"></p> | ||
| + | <center><em>Figure 2: 3D Absorbance graph for CPRG assay</em></center> | ||
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| + | <p><img src="https://static.igem.org/mediawiki/parts/1/11/2016_iGem_Toronto_GolS_2D.png" alt="alt text"></p> | ||
| + | <center><em>Figure 3: 2D Absorbance graph for CPRG assa</em></center> | ||
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| + | <p>We expected to see a linear increase correlated to increase in the concentration. However, instead, we observed a sudden decrease in expression levels after 0.5uM. This became more pronounced over time, resulting a Range Detector Circuit.</p> | ||
| + | <p>We suspect that there is an inherent edge that represses the expression after a certain threshold. This would produce the circuit shown in Figure 4. We speculated that this effect might be minimum for cell free extracts that contain minimal enzymes.</p> | ||
| + | <p><img src="https://static.igem.org/mediawiki/parts/6/60/Igem_2016_Circuit_Diagram.png" alt="alt text"></p> | ||
| + | <center><em>Figure 4: computational explanation for the CPRG assay</em></center> | ||
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<h3 id="not-so-successful">Not so successful</h3> | <h3 id="not-so-successful">Not so successful</h3> | ||
<ul> | <ul> | ||
| − | <li>Cell-free assay | + | <li>Cell-free assay required more time and troubleshooting.</li> |
</ul> | </ul> | ||
<h3 id="future-plans">Future plans</h3> | <h3 id="future-plans">Future plans</h3> | ||
<ul> | <ul> | ||
| − | <li>Make cell-free assays work</li> | + | <li>Make cell-free assays work.</li> |
</ul> | </ul> | ||
| − | < | + | <!--INGORE THIS STUFF, THIS FOR ME TO FINISH LATER WHEN I HAS ALL EXCEL FILES--> |
<p>The iGEM Toronto 2016 team was successful at modifying an naturally existing transcription factor, GolS (natively found in <em>Salmonella enterica</em>), to contain certain mutations that have been found to cause CueR (a copper-response factor in <em>Escherichia coli</em>)to increase sensitivity to gold and significantly decrease it's sensitivity to copper. This modification was due to an amino acid change from proline to alanine at site 118. This successful synthetic change can be found composite BioBricks, <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048002">Long GolS P118A (BBa_K2048002)</a> compared to the unchanged <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048001">Long GolS (BBa_K2048001)</a>.</p> | <p>The iGEM Toronto 2016 team was successful at modifying an naturally existing transcription factor, GolS (natively found in <em>Salmonella enterica</em>), to contain certain mutations that have been found to cause CueR (a copper-response factor in <em>Escherichia coli</em>)to increase sensitivity to gold and significantly decrease it's sensitivity to copper. This modification was due to an amino acid change from proline to alanine at site 118. This successful synthetic change can be found composite BioBricks, <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048002">Long GolS P118A (BBa_K2048002)</a> compared to the unchanged <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K2048001">Long GolS (BBa_K2048001)</a>.</p> | ||
| − | < | + | <!--These |
| − | </div></div><div id="tableofcontents" class="tableofcontents affix sidebar col-lg-4 hidden-xs hidden-sm hidden-md visible-lg-3"><ul class="nav" | + | Here you can describe the results of your project and your future plans. |
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| − | + | What should this page contain? | |
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| + | * Clearly and objectively describe the results of your work. | ||
| + | * Future plans for the project | ||
| + | * Considerations for replicating the experiments | ||
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| + | Project Achievements | ||
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| + | You can also include a list of bullet points (and links) of the successes and failures you have had over your summer. It is a quick reference page for the judges to see what you achieved during your summer. | ||
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| + | * A list of linked bullet points of the successful results during your project | ||
| + | * A list of linked bullet points of the unsuccessful results during your project. This is about being scientifically honest. If you worked on an area for a long time with no success, tell us so we know where you put your effort. | ||
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| + | Inspiration | ||
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| + | See how other teams presented their results. | ||
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| + | * [2014 TU Darmstadt](https://2014.igem.org/Team:TU_Darmstadt/Results/Pathway) | ||
| + | * [2014 Imperial](https://2014.igem.org/Team:Imperial/Results) | ||
| + | * [2014 Paris Bettencourt](https://2014.igem.org/Team:Paris_Bettencourt/Results) | ||
| + | --> | ||
| + | </div></div><div id="tableofcontents" class="tableofcontents affix sidebar col-lg-4 hidden-xs hidden-sm hidden-md visible-lg-3"><ul class="nav"> | ||
<li><a href="#success">Success</a></li> | <li><a href="#success">Success</a></li> | ||
| + | <li><a href="#discussion">Discussion</a></li> | ||
<li><a href="#not-so-successful">Not so successful</a></li> | <li><a href="#not-so-successful">Not so successful</a></li> | ||
<li><a href="#future-plans">Future plans</a></li> | <li><a href="#future-plans">Future plans</a></li> | ||
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</ul> | </ul> | ||
</div></div></div> | </div></div></div> | ||
Revision as of 10:27, 19 October 2016
Results
Success
- Both Long GolS P118A (BBa_K2048002) and Long GolS (BBa_K2048001) constructs responded to gold concentrations in a dose-responsive manner up to 1uM
Long GolS P118A (BBa_K2048002) is more responsive to gold than copper
Figure 1: CPRG assay results after 12 hours of incubation Long GolS (BBa_K2048001) is more responsive to both gold and copper concentrations than York 2013 construct (BBa_K1127008)
Discussion
We expected to see a linear increase correlated to increase in the concentration. However, instead, we observed a sudden decrease in expression levels after 0.5uM. This became more pronounced over time, resulting a Range Detector Circuit.
We suspect that there is an inherent edge that represses the expression after a certain threshold. This would produce the circuit shown in Figure 4. We speculated that this effect might be minimum for cell free extracts that contain minimal enzymes.
Not so successful
- Cell-free assay required more time and troubleshooting.
Future plans
- Make cell-free assays work.
The iGEM Toronto 2016 team was successful at modifying an naturally existing transcription factor, GolS (natively found in Salmonella enterica), to contain certain mutations that have been found to cause CueR (a copper-response factor in Escherichia coli)to increase sensitivity to gold and significantly decrease it's sensitivity to copper. This modification was due to an amino acid change from proline to alanine at site 118. This successful synthetic change can be found composite BioBricks, Long GolS P118A (BBa_K2048002) compared to the unchanged Long GolS (BBa_K2048001).
