http://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&feed=atom&action=historyTeam:St Andrews/project/modelling/model 3 - Revision history2024-03-29T06:32:51ZRevision history for this page on the wikiMediaWiki 1.16.5http://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=206108&oldid=prevAlly m: /* Conclusions */2010-10-28T02:51:07Z<p><span class="autocomment">Conclusions</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=Conclusions=</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=Conclusions=</div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">Our third modelling attempt could certainly be seen as the most successful and most useful for the purposes of our project. This model produced the switching behaviour we were looking for, and allowed us to perform detailed parameter tests on the system. What we have actually achieved is a tunable system, whereby the user can adjust the level of bistability to suit a particular need.</ins></div></td></tr>
</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=205956&oldid=prevAlly m: /* LuxR degradation */2010-10-28T02:47:00Z<p><span class="autocomment">LuxR degradation</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===LuxR degradation===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===LuxR degradation===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Similar tests were performed on the effects of changing the rate of LuxR degradation. We also expected interesting results from these tests since the LuxR is the HSL binding molecule. Our initial test to find a suitable parameter range gave us the data shown in figure .</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Similar tests were performed on the effects of changing the rate of LuxR degradation. We also expected interesting results from these tests since the LuxR is the HSL binding molecule. Our initial test to find a suitable parameter range gave us the data shown in figure <ins class="diffchange diffchange-inline">9</ins>.</div></td></tr>
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</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=205878&oldid=prevPatrickolden at 02:44, 28 October 20102010-10-28T02:44:30Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-26 16-52-06.jpg|center|800px]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-26 16-52-06.jpg|center|800px]]</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>'''Figure 10'''</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>'''Figure 10<ins class="diffchange diffchange-inline">: Point of up regulation as a function of RBS efficency and PoPs</ins>'''</div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">From this graphic we can see that, as predicted, the cell density at which the system turned on decreased as the PoPs and RBS efficiency were increased. The PoPs and the RBS efficency have almost the exact same effect on the point of up-regulation, hence the almost symmetrical plot. If viewed from above (as shown below) we can see that for very low PoPs & RBS efficiencies (<0.15) the system does not turn on at all.</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-26 16-52-30.jpg|center|800px]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-26 16-52-30.jpg|center|800px]]</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>'''Figure 11'''</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>'''Figure 11<ins class="diffchange diffchange-inline">: Point of up regulation as a function of RBS efficency and PoPs (from above)</ins>'''</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>'''Figure 12'''</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>'''Figure 12<ins class="diffchange diffchange-inline">: Delta cell density as a function of RBS efficeny and PoPs</ins>'''</div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">This plot confirms our initial hypothesis: changing the RBS expression and PoPs alters the "level of bistability" of the system. If analaysed (easier from above shown in the plot below) there is a very discernable blank spot at high PoPs and RBS efficencies. At this point the results are "Not A Number" which indicates that there is a malfuction in the model. This phonomena is explored further below.</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-26 16-48-54.jpg|center|800px]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-26 16-48-54.jpg|center|800px]]</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>'''Figure 13'''</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>'''Figure 13<ins class="diffchange diffchange-inline">: Delta cell density as a function of RBS efficeny and PoPs (from above)</ins>'''</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-28 01-52-10.png|800px]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Snapshot 2010-10-28 01-52-10.png|800px]]</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>'''Figure 14'''</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>'''Figure 14<ins class="diffchange diffchange-inline">: HSL, HSL-LuxR, LuxR Concentration for "Haywire" cell (High PoPs & RBS efficency)</ins>'''</div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> </div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">There are three phases in this graphic. The intial section shows the concentrations gradually increasing as the cell density is increased. Having switched on, the system then pleatues as predicted in low PoPs & RBS efficency models. However, in contrast to these models, the plateau then switches away from the steady state concentrations and reaches exponential values. Eventually this becomes a timestep issue, however further study is needed to understand why this happens intiially. Regardless, this happens at an extremely high cell density which in reality is never reached, and therefore our model should be giving realistic results when not in this parameter space. </ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=Conclusions=</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=Conclusions=</div></td></tr>
</table>Patrickoldenhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=203191&oldid=prevAlly m: /* Promoter efficiency and affinity */2010-10-28T00:58:31Z<p><span class="autocomment">Promoter efficiency and affinity</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Figure 13'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Figure 13'''</div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">'''Figure 14'''</ins></div></td></tr>
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</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=201571&oldid=prevAlly m: /* LuxR degradation */2010-10-27T23:58:42Z<p><span class="autocomment">LuxR degradation</span></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>There is a clear drop to zero in the bistability at a certain threshold level for LuxR degradation. This is due to the fact that the LuxR is degrading <del class="diffchange diffchange-inline">before an can bind to </del>the HSL <del class="diffchange diffchange-inline">so </del>the system <del class="diffchange diffchange-inline">does not switch </del>on.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>There is a clear drop to zero in the bistability at a certain threshold level for LuxR degradation. This is due to the fact that the LuxR is degrading <ins class="diffchange diffchange-inline">at such a high rate that there is not enough binding between it and </ins>the HSL <ins class="diffchange diffchange-inline">to sustain the quorum sensing network, and thus </ins>the system <ins class="diffchange diffchange-inline">never switches </ins>on <ins class="diffchange diffchange-inline">during the cell growth cycle. Before this point there is an exponential relationship between the level of bistability and the LuxR degradation rate. The explanation for this we hypothesise is the same as was the case for the HSL degradation, in that if the degradation rate is high the system takes longer to reach the equilibrium and so the bistability is greater</ins>.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Promoter efficiency and affinity===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Promoter efficiency and affinity===</div></td></tr>
</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=200708&oldid=prevAlly m: /* Implementation */2010-10-27T23:28:50Z<p><span class="autocomment">Implementation</span></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Our test indicated that our parameter test should be performed for values, 0.05 < kConvHSL < 0.25 and 0.0004 < kDegHSL < 0.0005. The next stage of testing was to run our simulation through these two ranges of values and observe the values produced for ΔCell density. The results of these tests are shown below. From these plots we can clearly see that by increasing the HSL conversion rate we also decrease the bistability of the system and by increasing the HSL degradation rate we increase the bistability of the system. Thinking through these two results, logical conclusions can be drawn. <del class="diffchange diffchange-inline">If kConv is </del>high, the <del class="diffchange diffchange-inline">system will generate large amounts </del>of HSL <del class="diffchange diffchange-inline">during the transition from OFF to ON</del>, and so when <del class="diffchange diffchange-inline">it comes to </del>the <del class="diffchange diffchange-inline">transition from ON to OFF</del>, <del class="diffchange diffchange-inline">there </del>is <del class="diffchange diffchange-inline">still </del>a <del class="diffchange diffchange-inline">high concentration </del>of <del class="diffchange diffchange-inline">HSL present in </del>the <del class="diffchange diffchange-inline">environment</del>. <del class="diffchange diffchange-inline">Hence </del>the system <del class="diffchange diffchange-inline">maintains </del>the <del class="diffchange diffchange-inline">same </del>steady state <del class="diffchange diffchange-inline">until this HSL supply diminishes</del>. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Our test indicated that our parameter test should be performed for values, 0.05 < kConvHSL < 0.25 and 0.0004 < kDegHSL < 0.0005. The next stage of testing was to run our simulation through these two ranges of values and observe the values produced for ΔCell density. The results of these tests are shown below. From these plots we can clearly see that by increasing the HSL conversion rate we also decrease the bistability of the system and by increasing the HSL degradation rate we increase the bistability of the system. Thinking through these two results, logical conclusions can be drawn. <ins class="diffchange diffchange-inline">With a </ins>high <ins class="diffchange diffchange-inline">value for kConv</ins>, the <ins class="diffchange diffchange-inline">saturation concentration </ins>of HSL <ins class="diffchange diffchange-inline">is reached much quicker</ins>, and so when the <ins class="diffchange diffchange-inline">cell death cycle is invoked</ins>, <ins class="diffchange diffchange-inline">the difference between the two graphs </ins>is <ins class="diffchange diffchange-inline">smaller than if </ins>a <ins class="diffchange diffchange-inline">low value had been used. The key point to think about is that kConv does not change the final concentrations of any </ins>of the <ins class="diffchange diffchange-inline">chemicals, rather it decreases the cell density required to reach those final concentrations</ins>.</div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">Looking now to the HSL degradation rate, a higher degradation rate will have the opposite effect of a high conversion rate, causing </ins>the system <ins class="diffchange diffchange-inline">to take a longer time to reach </ins>the steady state<ins class="diffchange diffchange-inline">. Thus when the cell death cycle is invoked a greater level of bistability will be displayed</ins>. </div></td></tr>
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</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=199190&oldid=prevAlly m: /* HSL production & degradation */2010-10-27T22:44:16Z<p><span class="autocomment">HSL production & degradation</span></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Our test indicated that our parameter test should be performed for values, 0.05 < kConvHSL < 0.25 and 0.0004 < kDegHSL < 0.0005. The results of these tests are shown below.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Our test indicated that our parameter test should be performed for values, 0.05 < kConvHSL < 0.25 and 0.0004 < kDegHSL < 0.0005<ins class="diffchange diffchange-inline">. The next stage of testing was to run our simulation through these two ranges of values and observe the values produced for ΔCell density</ins>. The results of these tests are shown below<ins class="diffchange diffchange-inline">. From these plots we can clearly see that by increasing the HSL conversion rate we also decrease the bistability of the system and by increasing the HSL degradation rate we increase the bistability of the system. Thinking through these two results, logical conclusions can be drawn. If kConv is high, the system will generate large amounts of HSL during the transition from OFF to ON, and so when it comes to the transition from ON to OFF, there is still a high concentration of HSL present in the environment. Hence the system maintains the same steady state until this HSL supply diminishes</ins>. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Figure 8: Parameter sensitivity test on rate of HSL degradation'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Figure 8: Parameter sensitivity test on rate of HSL degradation'''</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">From these plots we can clearly see that by increasing the HSL conversion rate we also decrease the bistability of the system and by increasing the HSL degradation rate we increase the bistability of the system.</del></div></td><td colspan="2"> </td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===LuxR degradation===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===LuxR degradation===</div></td></tr>
</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=198883&oldid=prevAlly m: /* Parameter Testing */2010-10-27T22:26:12Z<p><span class="autocomment">Parameter Testing</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===HSL production & degradation===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===HSL production & degradation===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We reasoned that the rate at which HSL was both produced, and degraded within the system should have a significant effect on the bistability since this molecule is fundamental to the process of quorum sensing in E.coli. Indeed our initial tests found that if the conversion rate was too low, or the degradation rate too high, the system did not actually switch on (see figure 6).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We reasoned that the rate at which HSL was both produced, and degraded within the system should have a significant effect on the bistability since this molecule is fundamental to the process of quorum sensing in E.coli. Indeed our initial tests found that if the conversion rate was too low, or the degradation rate too high, the system did not actually switch on (see figure 6)<ins class="diffchange diffchange-inline">. This would make biological sense, since if we have HSL being produced in the cells at a high rate there will be a plentiful supply with which the LuxR can bind. Similarly a low production rate or high degradation rate will result in a shortage of HSL</ins>.</div></td></tr>
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</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=198591&oldid=prevAlly m: /* Parameter Testing */2010-10-27T22:13:50Z<p><span class="autocomment">Parameter Testing</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Association and dissociation of HSL-LuxR===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Association and dissociation of HSL-LuxR===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>The HSL-LuxR complex is key to the entire quorum sensing circuit and as such we would expect it to play an important role in the level of bistability of the system. Our initial tests showed unsurprising results, that at high dissociation rates the system requires a higher rate of association to switch on and vice-versa. This is what we would expect, since when the association rate is high HSL-LuxR will accumulate much faster and so promote transcription much faster also. Similarly a high dissociation rate will result in very few HSL-LuxR molecules present to promote transcription and so GFP production will be less pronounced.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>The HSL-LuxR complex is key to the entire quorum sensing circuit and as such we would expect it to play an important role in the level of bistability of the system. Our initial tests showed unsurprising results, that at high dissociation rates the system requires a higher rate of association to switch on and vice-versa. This is what we would expect, since when the association rate is high HSL-LuxR will accumulate much faster and so promote transcription much faster also. Similarly a high dissociation rate will result in very few HSL-LuxR molecules present to promote transcription and so GFP production will be less pronounced<ins class="diffchange diffchange-inline">. There is a definite central plateau on the graph in which the rates of association and dissociation are such that the system switches on and this is the parameter space in which we were to perform our tests</ins>.</div></td></tr>
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</table>Ally mhttp://2010.igem.org/wiki/index.php?title=Team:St_Andrews/project/modelling/model_3&diff=198046&oldid=prevAlly m: /* Parameter Testing */2010-10-27T21:54:26Z<p><span class="autocomment">Parameter Testing</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Association and dissociation of HSL-LuxR===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Association and dissociation of HSL-LuxR===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>The HSL-LuxR complex is key to the entire quorum sensing circuit and as such we would expect it to play an important role in the level of bistability of the system. Our initial tests showed unsurprising results, that at high dissociation rates the system requires a higher rate of association to switch on and vice-versa.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>The HSL-LuxR complex is key to the entire quorum sensing circuit and as such we would expect it to play an important role in the level of bistability of the system. Our initial tests showed unsurprising results, that at high dissociation rates the system requires a higher rate of association to switch on and vice-versa<ins class="diffchange diffchange-inline">. This is what we would expect, since when the association rate is high HSL-LuxR will accumulate much faster and so promote transcription much faster also. Similarly a high dissociation rate will result in very few HSL-LuxR molecules present to promote transcription and so GFP production will be less pronounced</ins>.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===HSL production & degradation===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===HSL production & degradation===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We reasoned that the rate at which HSL was both produced, and degraded within the system should have a significant effect on the bistability since this molecule is fundamental to the process of quorum sensing in E.coli. Indeed our initial tests found that if the conversion rate was too low, or the degradation rate too high, the system did not actually switch on (see figure ).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We reasoned that the rate at which HSL was both produced, and degraded within the system should have a significant effect on the bistability since this molecule is fundamental to the process of quorum sensing in E.coli. Indeed our initial tests found that if the conversion rate was too low, or the degradation rate too high, the system did not actually switch on (see figure <ins class="diffchange diffchange-inline">6</ins>).</div></td></tr>
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</table>Ally m