|
|
| Line 134: |
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| | </tr> | | </tr> |
| | </table> | | </table> |
| - | | + | |
| - |
| + | <p> |
| - | | + | Working with the aTc/TetR parameters, we needed to now if our system could work with mercury. |
| | + | In order to see if it's <a href="https://2011.igem.org/Team:Grenoble/Projet/Results/Sensitivity#Mercury">applicable to mercury</a>, |
| | + | we realized a <a href="https://2011.igem.org/Team:Grenoble/Projet/Results/Sensitivity#Robustness">sensitivity to parameters study</a>. |
| | + | </p> |
| | | | |
| | | | |
| Line 165: |
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| | | | |
| | </div> | | </div> |
| - | <div class="blocbackground" id="Sensitivity">
| |
| - | <h2>Parameters sensitivity</h2>
| |
| - | <p>
| |
| - | In order to know if an error on the parameters would induce a completely different behaviour of our system,
| |
| - | we studied the sensitivity of our system to changes in the parameter values.
| |
| - | </p>
| |
| - | <p>
| |
| - | This study was performed by increasing or decreasing our parameters values by a range of percentages (from -66% to +300% for each parameter).
| |
| - | Then we studied the change on the output of our system, the ratio of IPTG over aTc that induces the coloration.
| |
| - | </p>
| |
| - | <p>
| |
| - | On the following figure one can see the influence on the output of our system (the ratio of IPTG over aTc on the interface)
| |
| - | for several aTc concentrations. For these concentrations the switch is still efficient, even though the resulting
| |
| - | variation on the output will induce an error on our measure.
| |
| - | </p>
| |
| - | <table class="nobordure">
| |
| - | <tr>
| |
| - | <td>
| |
| - | <center>
| |
| - | <a href= "https://static.igem.org/mediawiki/2011/4/41/Parameters_sensitivity1.png"><img src="https://static.igem.org/mediawiki/2011/4/41/Parameters_sensitivity1.png" style="width: 600px;" class="centerwide"/></a>
| |
| - | <div class="legend"><strong>Figure 1: </strong>Parameters sensitivity for 2.8e-07 M of aTc</div>
| |
| - | </center>
| |
| - | </td>
| |
| - | <td>
| |
| - | <p><strong>Note:</strong> On the next two figures the parameters are the following<br/>
| |
| - | <table style="border: 1px;">
| |
| - | <tr>
| |
| - | <td><strong>1</strong></td>
| |
| - | <td>k<SUB>plac</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>2</strong></td>
| |
| - | <td>V<SUB>cell</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>3</strong></td>
| |
| - | <td>k<SUB>pTet <br/></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>4</strong></td>
| |
| - | <td>K<SUB>pLac - LacI</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>5</strong></td>
| |
| - | <td>K<SUB>pTet - TerR</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>6</strong></td>
| |
| - | <td>K<SUB>LacI - IPTG</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>7</strong></td>
| |
| - | <td>K<SUB>TetR - aTc</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>8</strong></td>
| |
| - | <td>n<SUB>plac</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>9</strong></td>
| |
| - | <td>n<SUB>pTet</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>10</strong></td>
| |
| - | <td>δ<SUB>TetR</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>11</strong></td>
| |
| - | <td>δ<SUB>LacI</SUB></td>
| |
| - | </tr>
| |
| - |
| |
| - |
| |
| - | </table>
| |
| - | </td>
| |
| - | </tr>
| |
| - | </table>
| |
| - | <table class="nobordure">
| |
| - | <tr>
| |
| - | <td>
| |
| - | <a href="https://static.igem.org/mediawiki/2011/5/5b/Parameters_sensitivity2.png"><img src="https://static.igem.org/mediawiki/2011/5/5b/Parameters_sensitivity2.png" class ="centerwide" style="width: 600px;"/></a>
| |
| - | <div class="legend"><strong>Figure 2: </strong>Parameters sensitivity for 6.3e-06 M of aTc</div>
| |
| - | </td>
| |
| - | <td>
| |
| - | <p>Variations from 1 to 13 : </p>
| |
| - | <table>
| |
| - | <tr>
| |
| - | <td><strong>1</strong></td>
| |
| - | <td>-66%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>2</strong></td>
| |
| - | <td>-50%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>3</strong></td>
| |
| - | <td>-20%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>4</strong></td>
| |
| - | <td>-10%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>5</strong></td>
| |
| - | <td>-5%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>6</strong></td>
| |
| - | <td>+0%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>7</strong></td>
| |
| - | <td>+5%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>8</strong></td>
| |
| - | <td>+10%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>9</strong></td>
| |
| - | <td>+20%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>10</strong></td>
| |
| - | <td>+50%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>11</strong></td>
| |
| - | <td>+100%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>12</strong></td>
| |
| - | <td>+200%</td>
| |
| - | </tr>
| |
| - |
| |
| - | <tr>
| |
| - | <td><strong>13</strong></td>
| |
| - | <td>+300%</td>
| |
| - | </tr>
| |
| - | </table>
| |
| - | </td>
| |
| - | </tr>
| |
| - | </table>
| |
| - |
| |
| - | <p>For low values of aTc concentration the error is too minor to perturb the mechanism of our system. However,
| |
| - | it becomes impossible to predict the system output if the error is superior to -50% or +100% for parameters such
| |
| - | as K<SUB>pLac - LacI</SUB> or K<SUB>pTet - TetR</SUB>. In such a case a characterization of the responsible
| |
| - | parameter would be necessary.
| |
| - | </p>
| |
| - | <p>
| |
| - | For higher aTc concentrations however, the level of IPTG at which the switch occurs is very high. Errors on the parameters
| |
| - | prevents the system to switch for the chosen IPTG gradient. In this case, the IPTG maximal necessary valuefor quantification
| |
| - | would be too high for a living cell. The only problem being a decrease of the maximum value we can quantify.
| |
| - | </p>
| |
| - | <center>
| |
| - | <a href="https://static.igem.org/mediawiki/2011/0/01/Parameters_sensitivity3.png"><img src="https://static.igem.org/mediawiki/2011/0/01/Parameters_sensitivity3.png" class="centerwide" style="width: 850px;"/></a>
| |
| - | <div class="legend"><strong>Figure 2: </strong>Parameters sensitivity for 6.2e-04 M of aTc</div>
| |
| - | </center>
| |
| - |
| |
| - | </div>
| |
| - |
| |
| - |
| |
| - |
| |
| | <div id="selector"> | | <div id="selector"> |
| | <form method="get" > | | <form method="get" > |
| Line 385: |
Line 205: |
| | | | |
| | <option value="/Parameters" selected="selected">Our parameters</option> | | <option value="/Parameters" selected="selected">Our parameters</option> |
| - |
| |
| - | <option value="/Parameters#Sensitivity">Parameters sensitivity</option>
| |
| | | | |
| | </optgroup> | | </optgroup> |
Modelling - Parameters
Parameters
PDF file (detailed explanation on parameters and how we deduced them) for Toggle Switch
| kplac |
pLac production rate |
600 proteins/min |
[1] |
| kpTet |
pTet production rate |
600 proteins/min |
[1] |
| kpMer |
pMer production rate |
600 proteins/min |
[1] |
| δLacI |
LacI degradation rate |
4.6E-2 min−1 |
PDF file |
| δTetR |
TetR degradation rate |
4.6E-2 min−1 |
PDF file |
| δMerR |
MerR degradation rate |
4.6E-2 min−1 |
PDF file |
| KpLac - LacI |
plac - LacI dissociation constant |
5.45E-7 M |
Kyoto 10 |
| KpMer - MerR |
pMer - MerR dissociation constant |
1.00E-8 M |
[2] |
| KpTet - TerR |
pTet - TetR dissociation constant |
5.00E-8 M |
[3] |
| KLacI - IPTG |
LacI - IPTG dissociation constant |
2.96E-5 M |
[4] |
| KTetR - aTc |
aTc - TetR dissociation constant |
1.5E-8 M |
[5] |
| KHg2+ - MerR |
Hg2+ - MerR dissociation constant |
1E-7 M |
[2] |
| nplac |
plac cooperativity number |
2,3 |
[4] |
| npMer |
pMer cooperativity number |
2,5 |
[6] |
| npTet |
pTet cooperativity number |
3 |
[7] |
Working with the aTc/TetR parameters, we needed to now if our system could work with mercury.
In order to see if it's applicable to mercury,
we realized a sensitivity to parameters study.
References
[1] Nature. 2000 Jan 20;403(6767):335-8. A synthetic oscillatory network of transcriptional regulators. Elowitz MB, Leibler S.
[2] Mol Gen Genet. 1999 Aug;262(1):154-62. Purification and characterization of MerR, the regulator of the broad-spectrum mercury resistance genes in Streptomyces lividans 1326. Rother D, Mattes R, Altenbuchner J.
[3] David Braun et al. Parameter estimation for two synthetic gene networks: A case study. IEEE 2005.
[4] Nature 403, 339-342 (20 January 2000) Construction of a genetic toggle switch in Escherichia coli. Timothy S. Gardner Charles R. Cantor & James J. Collins
[5] O. Scholz, P. Schubert, M. Kintrup and W. Hillen, Tet repressor induction without Mg2+, Biochemistry 39 (2000), pp. 10914–10920
[6] Ultrasensitivity and heavy-metal selectivity of the allosterically modulated MerR transcription complex. D M Ralston and T V O'Halloran
[7] Systems analysis of a quorum sensing network: design constraints imposed by the functional requirements, network topology and kinetic constants. Goryachev AB, Toh DJ, Lee T.
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