Difference between revisions of "Part:BBa C0060:Experience"

Revision as of 14:16, 23 September 2011

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Applications of BBa_C0060

User Reviews

UNIQ6c83084d4a297aae-partinfo-00000000-QINU

•••••

UNIPV-Pavia iGEM 2011

NB: unless differently specified, all tests were performed in E. coli MGZ1 in M9 supplemented medium at 37°C in low copy plasmid pSB4C5.

The AiiA enzyme activity has been characterized under the regulation of p_tet promoter, assaying its enzymatic activity, with these parts:

BBa_K516220 p_Tet-RBS30-AiiA-TT
BBa_K516221 p_Tet-RBS31-AiiA-TT
BBa_K516222 p_Tet-RBS32-AiiA-TT
BBa_K516224 p_Tet-RBS34-AiiA-TT

A system of differential equations has been derived and we tried to identify the parameters, studying the exponential growth phase:

The four measurment parts ptet-RBSx-LuxI were quantitatively characetrized using the BBa_T9002 biosensor. Unfortunately we were not able to estimate the model parameters (k_cat, k_M,AiiA) because placing the device in low copy plasmid (pSB4C5) we did not observe HSL degradation, as you can see in the figures below:

We were only able to give semi-quantitative information about AiiA enzyme activity, performing an experiment in E. coli TOP10, with the four measurement devices in pSB1A2; the results are summerized in the graphs below:

In this way we can give a measure of AiiA activity as the percentage of HSL degraded in 7 hours (measurements provided as average [stdandard deviation]):

Measurement device	HSL degraded percentage
BBa_K516220	96.11 [3.66]
BBa_K516221	77.60 [2.66]
BBa_K516222	50.66 [16.87]
BBa_K516224	100 [0]

In order to evaluate the RBSs efficiency, we compute the ratio between the percentage of the degraded HSL realtive to the measurement device with the standard RBS (BBa_B0034):

RBS

RBS efficiency

BBa_B0030

0.96

BBa_B0031

0.78

BBa_B0032

0.51

BBa_B0034

1

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-<I>UNIPV-Pavia iGEM 2011</I>
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 <br>
-A system of differential equations (referring to <a href='http://2011.igem.org/Team:UNIPV-Pavia/Project/Modelling#Equations_for_gene_networks'>model equations</a>) has been derived and we tried to identify the parameters.
+A system of differential equations has been derived and we tried to identify the parameters, studying the exponential growth phase:
+<div style='text-align:justify'>
+<div class="thumbinner" style="width: 850px;">
+<a href="https://static.igem.org/mediawiki/2011/1/11/Pc_aiia.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/1/11/Pc_aiia.jpg" class="thumbimage" height="70%" width="68%"></a>
+</div></div>
+<br>
+The four measurment parts ptet-RBSx-LuxI were quantitatively characetrized using the <a href="https://parts.igem.org/Part:BBa_T9002:Experience#t9002">BBa_T9002 biosensor</a>. Unfortunately we were not able to estimate the model parameters (k<sub>cat</sub>, k<sub>M,AiiA</sub>) because placing the device in low copy plasmid (<a href="https://parts.igem.org/Part:pSB4C5">pSB4C5</a>) we did not observe HSL degradation, as you can see in the figures below:
+<table width='100%'><tr><td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/6/69/Aiia_LC_B0030.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/6/69/Aiia_LC_B0030.jpg" class="thumbimage" width="100%"></a></div></div>
+</td>
+<td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/6/63/Aiia_LC_B0031.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/6/63/Aiia_LC_B0031.jpg" class="thumbimage" width="100%"></a></div></div>
+</td></tr>
+<tr><td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/f/f5/Aiia_LC_B0032.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/f/f5/Aiia_LC_B0032.jpg" class="thumbimage" width="100%"></a></div></div>
+</td>
+<td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/8/8d/Aiia_LC_B0034.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/8/8d/Aiia_LC_B0034.jpg" class="thumbimage" width="100%"></a></div></div>
+</td></tr>
+</table>
+We were only able to give semi-quantitative information about AiiA enzyme activity, performing an experiment in <em>E. coli</em> TOP10, with the four measurement devices in <a href="https://parts.igem.org/Part:pSB1A2">pSB1A2</a>; the results are summerized in the graphs below:
+<table width='100%'><tr><td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/6/6c/Aiia_HC_B0030.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/6/6c/Aiia_HC_B0030.jpg" class="thumbimage" width="100%"></a></div></div>
+</td>
+<td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/e/ec/Aiia_HC_B0031.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/e/ec/Aiia_HC_B0031.jpg" class="thumbimage" width="100%"></a></div></div>
+</td></tr>
+<tr><td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/b/b2/Aiia_HC_B0032.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/b/b2/Aiia_HC_B0032.jpg" class="thumbimage" width="100%"></a></div></div>
+</td>
+<td width='50%'>
+<div style="text-align:justify"><div class="thumbinner" width='100%'>
+<a href="https://static.igem.org/mediawiki/2011/d/d7/Aiia_HC_B0034.jpg">
+<img alt="" src="https://static.igem.org/mediawiki/2011/d/d7/Aiia_HC_B0034.jpg" class="thumbimage" width="100%"></a></div></div>
+</td></tr>
+</table>
+In this way we can give a measure of AiiA activity as the percentage of HSL degraded in 7 hours (measurements provided as average [stdandard deviation]):
+<div align="center">
+<table class="data">
+<tr><td class="row"><b>Measurement device</b></td><td><b>   	HSL degraded percentage </b></td></tr>
+<tr><td class="row">BBa_K516220 </td><td class="row">		96.11 [3.66]</td></tr>
+<tr><td class="row">BBa_K516221 </td><td class="row">		77.60 [2.66]</td></tr>
+<tr><td class="row">BBa_K516222 </td><td class="row">		50.66 [16.87]</td></tr>
+<tr><td class="row">BBa_K516224</td><td class="row">	100 [0]</td></tr>
+</table></div>
+<br>
+In order to evaluate the RBSs efficiency, we compute the ratio between the percentage of the degraded HSL realtive to the measurement device with the standard RBS (BBa_B0034):
+<tr><td class="row"><b>RBS</b></td><td><b>   	RBS efficiency</b></td></tr>
+<tr><td class="row">BBa_B0030 </td><td class="row">		0.96</td></tr>
+<tr><td class="row">BBa_B0031</td><td class="row">		0.78</td></tr>
+<tr><td class="row">BBa_B0032</td><td class="row">		0.51</td></tr>
+<tr><td class="row">BBa_B0034</td><td class="row">	1</td></tr>
+</table></div>