Difference between revisions of "Team:Manchester/Description"

 
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<br /><br />
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<h1 class="title11">Project Overview </h1>
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<br /><br />
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<div class="team90">
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  <left>
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  <h1 class="mechanismm"> Mechanism 1</h1>
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  <h1 class="mectitle">Cell Free System</h1>
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<img class="piccog" src="https://static.igem.org/mediawiki/2016/5/5e/Manchester_TobyWikiPhoto.jpg"></img>
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  <img class="width80" style="margin:auto" src="https://static.igem.org/mediawiki/2016/8/86/T--Manchester--mechanism1_overview.png" alt="Mechanism 2 overview diagram" />
 
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<h3>★ ALERT! </h3>
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<p>This page is used by the judges to evaluate your team for the<a href="https://2016.igem.org/Judging/Medals"> improve a previous part or project gold medal criterion</a>. </p>
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<p> Delete this box in order to be evaluated for this medal. See more information at <a href="https://2016.igem.org/Judging/Pages_for_Awards/Instructions"> Instructions for Pages for awards</a>.</p>
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</div>
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<div class="info1">
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<div class="column onethird_size">
  
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  <p style="font-size:18px;text-align:left">Enzymatic colourimetric assays are used to determine the concentration of a chemical in a solution by the conversion of a chromogen substrate into a coloured product. We have introduced a plasmid expressing recombinant Alcohol Oxidase 1 (<a href="http://parts.igem.org/Part:BBa_K2092000" target="_blank">AOx</a>) from <i>Pichia pastoris</i> into <i>Escherichia coli</i>  BL21 (DE3) strain that will then be used in the cell-free colorimetric system. This method involves the usage of AOx to oxidise ethanol, producing hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) as a by-product. H<sub>2</sub>O<sub>2</sub> is used as an oxidising agent by horseradish peroxidase (HRP) to convert ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) to produce the colour change <sup>[1]</sup>. 
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  </p>
  
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<p>Tell us about your project, describe what moves you and why this is something important for your team.</p>
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<br /><br />
  
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<center>
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  <div class="directlink"><a href="https://2016.igem.org/Team:Manchester/Description/mechanism1"><h1>Click here for more info </h1></a></div>
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</center>
  
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<h5>What should this page contain?</h5>
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<ul>
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<li> A clear and concise description of your project.</li>
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</div>
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<li>A detailed explanation of why your team chose to work on this particular project.</li>
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<li>References and sources to document your research.</li>
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<li>Use illustrations and other visual resources to explain your project.</li>
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</ul>
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</div>
  
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<div class="column full_size" >
 
  
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<h5>Advice on writing your Project Description</h5>
 
  
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<p>
 
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We encourage you to put up a lot of information and content on your wiki, but we also encourage you to include summaries as much as possible. If you think of the sections in your project description as the sections in a publication, you should try to be consist, accurate and unambiguous in your achievements.
 
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</p>
 
  
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<p>
 
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Judges like to read your wiki and know exactly what you have achieved. This is how you should think about these sections; from the point of view of the judge evaluating you at the end of the year.
 
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</p>
 
  
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<br /><br /><br />
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<div class="box1"></div>
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  <h1 class="mechanismm1" > Mechanism 2</h1>
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  <h1 class="mectitle1">Inducible Gene Switch</h1>
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  <br /><br />
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<div class="column twothird_size">
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<center>
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  <img class="width90" style="margin:auto" src="https://static.igem.org/mediawiki/2016/1/12/T--Manchester--mech2_overview.png" alt="Mechanism 2 overview diagram" />
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</center>
 
</div>
 
</div>
  
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<div class="info2">
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<div class="column onethird_size">
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  <p style="font-size: 18px;text-align:left">The <i>alc</i> gene expression system is one of the most reliable chemically inducible gene switches for use in plants <sup>[2]</sup> and fungus <sup>[3]</sup>.
 +
  This system relies on the ability of <a href="http://parts.igem.org/Part:BBa_K2092001" target="_blank"> AlcR</a>, an alcohol-activated transcription factor, to bind to its target <i>alcA</i> promoter (<a href="http://parts.igem.org/Part:BBa_K2092002" target="_blank">P<i>alc</i>A</a>). Based on this, we have engineered  <i>E. coli</i> K-12 derivative DH5α and BL21 to induce expression of chromoproteins when AlcR binds to the native P<i>alc</i>A and variant of P<i>alc</i>A (<a href="http://parts.igem.org/Part:BBa_K2092003" target="_blank"> P<i>alc</i>A(var)</a>) in the presence of ethanol <sup>[4]</sup>.
 +
  </p>
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    <br /><br />
  
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<div class="column half_size" >
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<center>
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  <div class="directlink"><a href="https://2016.igem.org/Team:Manchester/Description/mechanism2"><h1>Click here for more info </h1></a></div>
 +
</center>  
  
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<h5>References</h5>
 
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<p>iGEM teams are encouraged to record references you use during the course of your research. They should be posted somewhere on your wiki so that judges and other visitors can see how you thought about your project and what works inspired you.</p>
 
  
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</div>
 
</div>
 
</div>
  
  
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<div class="column half_size" >
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<!------------------------------------------------Reference--------------------------------------------------------->
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<h5>Inspiration</h5>
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<div class="box1"></div>
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<p>See how other teams have described and presented their projects: </p>
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<center>
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<div class="referencediv">
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  <h1 class="reference">References</h1>
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  <ul class="romanlist">
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      <li style="text-align:left;"> Azevedo, A. M., Prazeres, D. M. F., Cabral, J. M., & Fonseca, L. P. (2005). Ethanol biosensors based on alcohol oxidase. <i>Biosensors and Bioelectronics</i>,21(2), 235-247.
 +
    </li>
 +
<li style="text-align: left;;"> Plants: Kinkema, M., Geijskes, R.J., Shand, K., Coleman, H.D., De Lucca, P.C., Palupe, A., Harrison, M.D., Jepson, I., Dale, J.L. and Sainz, M.B. (2013). An improved chemically inducible gene switch that functions in the monocotyledonous plant sugar cane. Plant Molecular Biology, 84(4-5), 443–454.
 +
</li>
 +
    <li style="text-align: left;;"> Panozzo, C., Capuano, V., Fillinger, S. and Felenbok, B. (1997). The zinc binuclear cluster Activator AlcR is able to bind to single sites but requires multiple repeated sites for synergistic activation of the alcA gene in Aspergillus nidulans. <i>Journal of Biological Chemistry</i>, 272(36), pp. 22859–22865.
 +
    </li>
 +
<li style="text-align: left;;"> Garoosi, A.G., Salter,M.G. , Caddick ,X.M and Tomsett, M.B. (2004). Characterization of the ethanol-inducible <i>alc</i> gene expression system in tomato. <i>Journal of experimental Botany</i>, 46 (416), pp. 1635-1642.
 +
</li>
 +
  </ul>
 +
 
 +
</div>
 +
</center>
 +
 
 +
</div>
  
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<ul>
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<div class="floatleft1 project1">
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<li><a href="https://2014.igem.org/Team:Imperial/Project"> Imperial</a></li>
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<a class="projectlink" href="https://2016.igem.org/Team:Manchester"><< Main Page</a>
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<li><a href="https://2014.igem.org/Team:UC_Davis/Project_Overview"> UC Davis</a></li>
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<li><a href="https://2014.igem.org/Team:SYSU-Software/Overview">SYSU Software</a></li>
+
−
</ul>
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</div>
 
</div>
  

Latest revision as of 14:39, 24 November 2016

Manchester iGEM 2016


Project Overview



Mechanism 1

Cell Free System
Mechanism 2 overview diagram

Enzymatic colourimetric assays are used to determine the concentration of a chemical in a solution by the conversion of a chromogen substrate into a coloured product. We have introduced a plasmid expressing recombinant Alcohol Oxidase 1 (AOx) from Pichia pastoris into Escherichia coli BL21 (DE3) strain that will then be used in the cell-free colorimetric system. This method involves the usage of AOx to oxidise ethanol, producing hydrogen peroxide (H2O2) as a by-product. H2O2 is used as an oxidising agent by horseradish peroxidase (HRP) to convert ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) to produce the colour change [1].



Click here for more info




Mechanism 2

Inducible Gene Switch



Mechanism 2 overview diagram

The alc gene expression system is one of the most reliable chemically inducible gene switches for use in plants [2] and fungus [3]. This system relies on the ability of AlcR, an alcohol-activated transcription factor, to bind to its target alcA promoter (PalcA). Based on this, we have engineered E. coli K-12 derivative DH5α and BL21 to induce expression of chromoproteins when AlcR binds to the native PalcA and variant of PalcA ( PalcA(var)) in the presence of ethanol [4].



Click here for more info

References

  • Azevedo, A. M., Prazeres, D. M. F., Cabral, J. M., & Fonseca, L. P. (2005). Ethanol biosensors based on alcohol oxidase. Biosensors and Bioelectronics,21(2), 235-247.
  • Plants: Kinkema, M., Geijskes, R.J., Shand, K., Coleman, H.D., De Lucca, P.C., Palupe, A., Harrison, M.D., Jepson, I., Dale, J.L. and Sainz, M.B. (2013). An improved chemically inducible gene switch that functions in the monocotyledonous plant sugar cane. Plant Molecular Biology, 84(4-5), 443–454.
  • Panozzo, C., Capuano, V., Fillinger, S. and Felenbok, B. (1997). The zinc binuclear cluster Activator AlcR is able to bind to single sites but requires multiple repeated sites for synergistic activation of the alcA gene in Aspergillus nidulans. Journal of Biological Chemistry, 272(36), pp. 22859–22865.
  • Garoosi, A.G., Salter,M.G. , Caddick ,X.M and Tomsett, M.B. (2004). Characterization of the ethanol-inducible alc gene expression system in tomato. Journal of experimental Botany, 46 (416), pp. 1635-1642.
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