http://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&feed=atom&action=historyTeam:Minnesota/Parts Characterization - Revision history2024-03-29T11:34:26ZRevision history for this page on the wikiMediaWiki 1.16.5http://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=131944&oldid=prevYiannis at 18:51, 20 October 20092009-10-20T18:51:31Z<p></p>
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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;">Although we will be judged only for the software we developed, an important aspect of our work is the combination of experiments and simulations. For detailed comparisons please look at the Notebook in the end of July. </del></div></td><td colspan="2"> </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;">We are submitting our constructs to the Registry. </del></div></td><td colspan="2"> </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;">We also characterized a handful of older BioBricks. Here are the details we are entering in the Registry.</del></div></td><td colspan="2"> </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;"><h1>Comments</h1></ins></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;"></ins></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;">Although we will be judged only for the software we developed, an important aspect of our work is the combination of experiments and simulations. For detailed comparisons please look at the Notebook in the end of July. </ins></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;"></ins></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;">We are submitting our constructs to the Registry. </ins></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;"></ins></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;">We also characterized a handful of older BioBricks. Here are the details we are entering in the Registry.</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h1>Parts Characterization</h1></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h1>Parts Characterization</h1></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h2>Introduction</h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h2>Introduction</h2></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After an exhaustive search of the [http://www.partsregistry.org/Main_Page Registry of Standard Parts], we found five promoters that were included in the 2009 iGEM Kit as potential parts characterize. Using part [http://partsregistry.org/wiki/index.php/Part:BBa_F2620 BBa_F2620], which was characterized by a group from MIT in 2004, as a template, we examined the following parts:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After an exhaustive search of the [http://www.partsregistry.org/Main_Page Registry of Standard Parts], we found five promoters that were included in the 2009 iGEM Kit as potential parts characterize. Using part [http://partsregistry.org/wiki/index.php/Part:BBa_F2620 BBa_F2620], which was characterized by a group from MIT in 2004, as a template, we examined the following parts:</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>The second promoter we examined was [http://partsregistry.org/Part:BBa_J13002 J13002], which was submitted to the Registry by [<del class="diffchange diffchange-inline">http</del>://<del class="diffchange diffchange-inline">parts</del>.<del class="diffchange diffchange-inline">mit</del>.<del class="diffchange diffchange-inline">edu/wiki/index.php</del>/UT_Austin_2005 UT Austin] in 2005. The part consists of two TetR binding sites and RBS and is called a "TetR repressed PoPs/RIPS generator." Since the part already had a RBS, we attached [http://partsregistry.org/Part:BBa_E0040 GFP] in order to indirectly measure PoPS. The results of our characterization can be found on the [http://partsregistry.org/Part:BBa_J13002:Experience Experience] page at the Registry for Part J13002. A photo of this part from the [http://<del class="diffchange diffchange-inline">parts</del>.<del class="diffchange diffchange-inline">mit.edu</del>/ Registry of Standard Parts] is displayed below: [[Image:J13002.jpg|700px|left]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>The second promoter we examined was [http://partsregistry.org/Part:BBa_J13002 J13002], which was submitted to the Registry by [<ins class="diffchange diffchange-inline">https</ins>://<ins class="diffchange diffchange-inline">2006</ins>.<ins class="diffchange diffchange-inline">igem</ins>.<ins class="diffchange diffchange-inline">org</ins>/UT_Austin_2005 UT Austin] in 2005. The part consists of two TetR binding sites and RBS and is called a "TetR repressed PoPs/RIPS generator." Since the part already had a RBS, we attached [http://partsregistry.org/Part:BBa_E0040 GFP] in order to indirectly measure PoPS. The results of our characterization can be found on the [http://partsregistry.org/Part:BBa_J13002:Experience Experience] page at the Registry for Part J13002. A photo of this part from the [http://<ins class="diffchange diffchange-inline">partsregistry</ins>.<ins class="diffchange diffchange-inline">org</ins>/ Registry of Standard Parts] is displayed below: [[Image:J13002.jpg|700px|left]]</div></td></tr>
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</table>Yiannishttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=131877&oldid=prevYiannis at 18:46, 20 October 20092009-10-20T18:46:12Z<p></p>
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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;">Although we will be judged only for the software we developed, an important aspect of our work is the combination of experiments and simulations. For detailed comparisons please look at the Notebook in the end of July. </ins></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;"></ins></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;">We are submitting our constructs to the Registry. </ins></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;"></ins></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;">We also characterized a handful of older BioBricks. Here are the details we are entering in the Registry.</ins></div></td></tr>
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</table>Yiannishttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=41006&oldid=prevLinkri at 19:23, 7 August 20092009-08-07T19:23:34Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota|<font color="gold">Home</font>]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota|<font color="gold">Home</font>]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Team|<font color="gold">The Team</font>]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Team|<font color="gold">The Team</font>]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Project|<font color="gold">The Project</font>]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Project|<font color="gold">The Project</font>]]</div></td></tr>
<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;">!align="center"|[[Team:Minnesota/Parts|<font color="gold">Submitted Parts</font>]]</del></div></td><td colspan="2"> </td></tr>
<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;">!align="center"|[[Team:Minnesota/Modeling|<font color="gold">Modeling</font>]]</del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Designer|<font color="gold">SynBioSS Designer</font>]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Designer|<font color="gold">SynBioSS Designer</font>]]</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/<del class="diffchange diffchange-inline">Parts Characterization</del>|<font color="gold"><del class="diffchange diffchange-inline">Parts Characterization</del></font>]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/<ins class="diffchange diffchange-inline">Modeling</ins>|<font color="gold"><ins class="diffchange diffchange-inline">Modeling</ins></font>]]</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Notebook|<font color="gold"><del class="diffchange diffchange-inline">Experiments and Calendar</del></font>]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>!align="center"|[[Team:Minnesota/Notebook|<font color="gold"><ins class="diffchange diffchange-inline">Experimental</ins></font>]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h1>Parts Characterization</h1></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h1>Parts Characterization</h1></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h2>Introduction</h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h2>Introduction</h2></div></td></tr>
</table>Linkrihttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=40966&oldid=prevDeckera at 16:53, 7 August 20092009-08-07T16:53:58Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>As you can see on the figure below, for each timepoint, the production of GFP is virtually identical. This confirmed the hypothesis that the promoter is actually constituitive rather than repressible. Regardless of IPTG concentration, we saw the same amount of GFP production over all the timepoints. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>As you can see on the figure below, for each timepoint, the production of GFP is virtually identical. This confirmed the hypothesis that the promoter is actually constituitive rather than repressible. Regardless of IPTG concentration, we saw the same amount of GFP production over all the timepoints. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:<del class="diffchange diffchange-inline">IPTG_TF2</del>.jpg|700px|center|Transfer function for Part I14032]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:<ins class="diffchange diffchange-inline">IPTG_TF3</ins>.jpg |700px|center|Transfer function for Part I14032]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We also examined the GFP production at varying IPTG concentrations over multiple rounds of cell division. As you can see from the figure below, not only is the constituitive promoter reliably on, but it continues to produce GFP at the same rate regardless of the number of cell divisions. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We also examined the GFP production at varying IPTG concentrations over multiple rounds of cell division. As you can see from the figure below, not only is the constituitive promoter reliably on, but it continues to produce GFP at the same rate regardless of the number of cell divisions. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:<del class="diffchange diffchange-inline">IPTG_stab</del>.jpg|600px|center|Stability of Part I14032]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:<ins class="diffchange diffchange-inline">IPTG_stab3</ins>.jpg|600px|center|Stability of Part I14032]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td></tr>
</table>Deckerahttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=40963&oldid=prevDeckera at 16:44, 7 August 20092009-08-07T16:44:11Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In the Experience section on the Registry, it was stated that although this promoter was classified as repressible, it was actually constituitive. However, there were no data to support this statement so we decided to investigate the promoter. In addition to investigating the promoter inducibility status, we also wanted to examine Transfer Function and Stability. So we attached the [http://partsregistry.org/Part_Bba_K081012 PoPS generator] to this promoter in a ligation reaction and induced it at various concentrations of IPTG: 0, 0.001, 0.01, 0.05 and 1 mM. We took samples over 9 hours and analyzed the products using flow cytometry. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In the Experience section on the Registry, it was stated that although this promoter was classified as repressible, it was actually constituitive. However, there were no data to support this statement so we decided to investigate the promoter. In addition to investigating the promoter inducibility status, we also wanted to examine Transfer Function and Stability. So we attached the [http://partsregistry.org/Part_Bba_K081012 PoPS generator] to this promoter in a ligation reaction and induced it at various concentrations of IPTG: 0, 0.001, 0.01, 0.05 and 1 mM. We took samples over 9 hours and analyzed the products using flow cytometry. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image:IPTG_TF.jpg|700px|left]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As you can see on the figure <ins class="diffchange diffchange-inline">below</ins>, for each timepoint, the production of GFP is virtually identical. This confirmed the hypothesis that the promoter is actually constituitive rather than repressible. Regardless of IPTG concentration, we saw the same amount of GFP production over all the timepoints. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As you can see on the figure <del class="diffchange diffchange-inline">to the left</del>, for each timepoint, the production of GFP is virtually identical. This confirmed the hypothesis that the promoter is actually constituitive rather than repressible. Regardless of IPTG concentration, we saw the same amount of GFP production over all the timepoints. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">[[Image:IPTG_TF2.jpg|700px|center|Transfer function for Part I14032]]</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image:IPTG_stability.jpg|600px|right]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We also examined the GFP production at varying IPTG concentrations over multiple rounds of cell division. As you can see from the <ins class="diffchange diffchange-inline">figure below</ins>, not only is the constituitive promoter reliably on, but it continues to produce GFP at the same rate regardless of the number of cell divisions. </div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We also examined the GFP production at varying IPTG concentrations over multiple rounds of cell division. As you can see from the <del class="diffchange diffchange-inline">graphs on the right</del>, not only is the constituitive promoter reliably on, but it continues to produce GFP at the same rate regardless of the number of cell divisions. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We also wanted to examine Transfer Function and Stability in this promoter. To do this, we grew 1 ml liquid cultures in different concentration of inducer media: 0, 1, 10, 50, 100 and 200 ng/ml of aTc. As you can see in the figure below, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We also wanted to examine Transfer Function and Stability in this promoter. To do this, we grew 1 ml liquid cultures in different concentration of inducer media: 0, 1, 10, 50, 100 and 200 ng/ml of aTc. As you can see in the figure below, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:<del class="diffchange diffchange-inline">ATc_TF</del>.jpg|center|700px|Transfer function for part J13002]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:<ins class="diffchange diffchange-inline">ATC_TF</ins>.jpg |center|700px|Transfer function for part J13002]]</div></td></tr>
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</table>Deckerahttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=40916&oldid=prevDeckera at 15:56, 7 August 20092009-08-07T15:56:12Z<p></p>
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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;">We also wanted to examine Transfer Function and Stability in this promoter. To do this, we grew 1 ml liquid cultures in different concentration of inducer media: 0, 1, 10, 50, 100 and 200 ng/ml of aTc. </del></div></td><td colspan="2"> </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;"><b3 /></ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image</del>:<del class="diffchange diffchange-inline">Atc_TF</del>.<del class="diffchange diffchange-inline">jpg|left|700px]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">We also wanted to examine Transfer Function and Stability in this promoter. To do this, we grew 1 ml liquid cultures in different concentration of inducer media</ins>: <ins class="diffchange diffchange-inline">0, 1, 10, 50, 100 and 200 ng/ml of aTc</ins>. As you can see in the <ins class="diffchange diffchange-inline">figure below</ins>, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As you can see in the <del class="diffchange diffchange-inline">graphs to the left</del>, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">[[Image:ATc_TF.jpg|center|700px|Transfer function for part J13002]]</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><br /></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><br /></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image:Atc_stab2.jpg|right|500px]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We also examined the stability of the device, which is how the production of GFP changes for each inducer concentration over multiple rounds of cell division. As you can see from the <ins class="diffchange diffchange-inline">figure below</ins>, the device is clearly inducible with a general downward trend in GFP production over time. The graph of aTc concentration of 50 ng/ml again demonstrates the spike in GFP production that we observed at this concentration. This is the optimal inducer concentration for the device. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline"><br /></del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">[[Image:ATc_stab.jpg|center|500px|Stability of part J13002]]</ins></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We also examined the stability of the device, which is how the production of GFP changes for each inducer concentration over multiple rounds of cell division. As you can see from the <del class="diffchange diffchange-inline">graphs to the right</del>, the device is clearly inducible with a general downward trend in GFP production over time. The graph of aTc concentration of 50 ng/ml again demonstrates the spike in GFP production that we observed at this concentration. This is the optimal inducer concentration for the device. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Unfortunately, we ran out of time for characterization of the third and final part, [http://partsregistry.org/Part:BBa_K091101 K091101]. However, we did characterize two existing BioBrick parts in terms of Transfer Function and Stability. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Unfortunately, we ran out of time for characterization of the third and final part, [http://partsregistry.org/Part:BBa_K091101 K091101]. However, we did characterize two existing BioBrick parts in terms of Transfer Function and Stability. </div></td></tr>
</table>Deckerahttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=40889&oldid=prevDeckera at 15:47, 7 August 20092009-08-07T15:47:43Z<p></p>
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<td colspan='2' style="background-color: white; color:black;">← Older revision</td>
<td colspan='2' style="background-color: white; color:black;">Revision as of 15:47, 7 August 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:<del class="diffchange diffchange-inline">Atc_transfer_function1</del>.jpg|left|700px]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:<ins class="diffchange diffchange-inline">Atc_TF</ins>.jpg|left|700px]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>As you can see in the graphs to the left, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>As you can see in the graphs to the left, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Stability</h4></div></td></tr>
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</table>Deckerahttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=40852&oldid=prevDeckera at 15:34, 7 August 20092009-08-07T15:34:54Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3>Promoter J13002: aTc regulation</h3></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>The second promoter we examined was [http://partsregistry.org/Part:BBa_J13002 J13002], which was submitted to the Registry by [http://parts.mit.edu/wiki/index.php/UT_Austin_2005 UT Austin] in 2005. The part consists of two TetR binding sites and RBS and is called a "TetR repressed PoPs/RIPS generator." Since the part already had a RBS, we attached [http://partsregistry.org/Part:BBa_E0040 GFP] in order to indirectly measure PoPS. A photo of this part from the [http://parts.mit.edu/ Registry of Standard Parts] is displayed below: [[Image:J13002.jpg|700px|left]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>The second promoter we examined was [http://partsregistry.org/Part:BBa_J13002 J13002], which was submitted to the Registry by [http://parts.mit.edu/wiki/index.php/UT_Austin_2005 UT Austin] in 2005. The part consists of two TetR binding sites and RBS and is called a "TetR repressed PoPs/RIPS generator." Since the part already had a RBS, we attached [http://partsregistry.org/Part:BBa_E0040 GFP] in order to indirectly measure PoPS<ins class="diffchange diffchange-inline">. The results of our characterization can be found on the [http://partsregistry.org/Part:BBa_J13002:Experience Experience] page at the Registry for Part J13002</ins>. A photo of this part from the [http://parts.mit.edu/ Registry of Standard Parts] is displayed below: [[Image:J13002.jpg|700px|left]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Atc_transfer_function1.jpg|left|700px]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Atc_transfer_function1.jpg|left|700px]]</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As you can see in the graphs to the left, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction<del class="diffchange diffchange-inline">. The results of our characterization can be found on the [http://partsregistry.org/Part:BBa_J13002:Experience Experience] page at the Registry for Part J13002</del>. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As you can see in the graphs to the left, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td></tr>
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</table>Deckerahttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=40848&oldid=prevDeckera at 15:33, 7 August 20092009-08-07T15:33:26Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h4>Transfer Function</h4></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Atc_transfer_function1.jpg|left|700px]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Atc_transfer_function1.jpg|left|700px]]</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As you can see in the graphs to the left, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As you can see in the graphs to the left, there was a general upward trend of GFP production with the concentration of aTc. The error bars in the graphs represent the standard error. Interestingly, we observed a spike in GFP production at 50 ng/ml of aTc. During the sampling, the culture growing in 50 ng/ml aTc inducer media did not grow as fast as the others, with the OD<sub>595</sub> as low as 0.08 rather than the preferred 0.2. The cells were putting their energy towards GFP production rather than growing, so the lack of growth and spike of mean GFP is legitimate. Since the inducer is an antibiotic, we hypothesized that at higher concentrations of 100 ng/ml and 200 ng/ml, the cells were using all their energy dealing with the aTc rather than GFP production. We did perform this experiment again but did not see any induction<ins class="diffchange diffchange-inline">. The results of our characterization can be found on the [http://partsregistry.org/Part:BBa_J13002:Experience Experience] page at the Registry for Part J13002</ins>. </div></td></tr>
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</table>Deckerahttp://2009.igem.org/wiki/index.php?title=Team:Minnesota/Parts_Characterization&diff=39844&oldid=prevDeckera at 15:50, 6 August 20092009-08-06T15:50:08Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/6/68/Restriction_Digest.pdf Digest] the promoter, GFP and plasmid backbone with the appropriate restriction enzymes for later ligation</li></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/6/68/Restriction_Digest.pdf Digest] the promoter, GFP and plasmid backbone with the appropriate restriction enzymes for later ligation</li></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/a/a8/DNA_Purification.pdf Gel purify] digest products</li></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/a/a8/DNA_Purification.pdf Gel purify] digest products</li></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/b/b3/DNA_Fragment_Ligation.pdf Ligate] the promoter + RBS and GFP after restriction enzyme digest in the plasmid backbone pSB3K3 overnight<li></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/b/b3/DNA_Fragment_Ligation.pdf Ligate] the promoter + RBS and GFP after restriction enzyme digest in the plasmid backbone pSB3K3 overnight<<ins class="diffchange diffchange-inline">/</ins>li></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/b/b0/Sequencing.pdf Sequence] ligation products<li></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/b/b0/Sequencing.pdf Sequence] ligation products<<ins class="diffchange diffchange-inline">/</ins>li></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/f/f5/Transformation_of_Chemically_Competent_Cells.pdf Transform] the ligation products into DH5alphaPro cells, which express TetR</li></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>[https://static.igem.org/mediawiki/2009/f/f5/Transformation_of_Chemically_Competent_Cells.pdf Transform] the ligation products into DH5alphaPro cells, which express TetR</li></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>Pick colonies and grow in 2ml liquid cultures plus kanamycin</li></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li>Pick colonies and grow in 2ml liquid cultures plus kanamycin</li></div></td></tr>
</table>Deckera