Team:Cornell/testing/project/wetlab/4/7

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<h2 class="centered">Reactor Testing</h2>
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<tr><th style="width:8px" class="header"></th><th style="width:8px" class="header"></th><th style="width:8px" class="header"></th><th style="width: 87px;" class="header">Name</th><th style="width: 80px" colspan="" class="header">Type</th><th style="width: auto;" colspan="" class="header">Description</th><th style="width: 150px" colspan="" class="header">Designer</th><th style="width: 50px" colspan="" class="header">Length</th>
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<td class="status_cell heart13">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957000">BBa_K957000</a></td><td>Generator</td><td>Arsenic inducible mtrB with cut sites flanking RBS</td><td width="100px">Dylan Patrick Webster</td><td align="right">2625</td>
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<td class="status_cell heart13">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957001">BBa_K957001</a></td><td>Generator</td><td>Arsenic inducible mtrB</td><td width="100px">Dylan Patrick Webster</td><td align="right">2704</td>
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<td class="status_cell heart13">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957002">BBa_K957002</a></td><td>Generator</td><td>Salicylate inducible mtrB with cut sites flanking RBS </td><td width="100px">Dylan Patrick Webster</td><td align="right">3207</td>
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<td class="status_cell heart13">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957003">BBa_K957003</a></td><td>Generator</td><td>Salicylate inducible mtrB</td><td width="100px">Dylan Patrick Webster</td><td align="right">3284</td>
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<td class="status_cell heart13">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957004">BBa_K957004</a></td><td>Coding</td><td>nah Operon for degradation of naphthalene to salicylate</td><td width="100px">Dylan Patrick Webster</td><td align="right">9409</td>
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<td class="status_cell heart13">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957005">BBa_K957005</a></td><td>Generator</td><td> Constitutive expression of nah Operon for degradation of naphthalene to salicylate </td><td width="100px">Dylan Patrick Webster</td><td align="right">9452</td>
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<td class="status_cell  "></td><td class="status_cell cell_white">&nbsp;</td><td class="status_cell cell_white">&nbsp;</td><td><a class="noul_link" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K957006">BBa_K957006</a></td><td>Composite</td><td>Constitutive expression of nah Operon with oriT </td><td width="100px">Dylan Patrick Webster</td><td align="right">9831</td>
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<h3>Overview of Characterization in Bioelectrochemical Stystems</h3>
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As described in the <a href="https://2012.igem.org/Team:Cornell/testing/project/wetlab/2">Chassis</a> section, <i>S. oneidensis</i> MR-1 is capable of shuttling electrons through the Mtr pathway to reduce Fe(III) to Fe(II) because of the negative free energy change associated with the redox reaction. Thus, to encourage <i>Shewanella</i> to transfer electrons to an electrode, we poise the redox potential of an electrode to that of Fe(III)/Fe(II) in a three electrode system, controlled by a potentiostat. [[LINK TO THE PAGE ON THE POTENTIOSTAT]]. In short, this instrument works by setting the potential of a working electrode (WE) with respect to a Ag/AgCl reference eletrode (RE) by injecting current through a counter electrode (CE). These electrodes can be seen in the schematic representation of our single-compartment bioelectrochemical reactors shown below.
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Revision as of 03:30, 4 October 2012

NameTypeDescriptionDesignerLength
    BBa_K957000GeneratorArsenic inducible mtrB with cut sites flanking RBSDylan Patrick Webster2625
    BBa_K957001GeneratorArsenic inducible mtrBDylan Patrick Webster2704
    BBa_K957002GeneratorSalicylate inducible mtrB with cut sites flanking RBS Dylan Patrick Webster3207
    BBa_K957003GeneratorSalicylate inducible mtrBDylan Patrick Webster3284
    BBa_K957004Codingnah Operon for degradation of naphthalene to salicylateDylan Patrick Webster9409
    BBa_K957005Generator Constitutive expression of nah Operon for degradation of naphthalene to salicylate Dylan Patrick Webster9452
  BBa_K957006CompositeConstitutive expression of nah Operon with oriT Dylan Patrick Webster9831
iGEM012 Cornell

First Lessons Learned: Control Reactors

In order to enhance the field-deployability of our final device, we initially decided to feed our reactors with LB, since a very concentrated LB source fed at a low flow rate could sustain our field reactors for extended periods of time without taking up much physical space. However, upon setting up control reactors—both in batch and continuous flow operation—we discovered that wild type S. oneidensis MR-1 produced significantly less current when fed with LB than M4—a commonly used media for Shewanella-inoculated bioelectrochemical systems.

Naphthalene & Salicylate Sensing

First characterized naphthalene sensors with salicylate, since our system indirectly senses naphthalene via salicylate.
Uninduced salicylate reporter produces same current as MR-1 in rich M4 media.
First attempt to diagnose problem: Switch to minimal M4... Producing higher current, but we're still not getting a salicylate response.
Next attempt: Poise the working electrode at a higher potential in attempt to get more MtrC and MtrA for MtrB to associate with. Didn't quite do the trick either.

Arsenic Sensing

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