http://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&feed=atom&action=historyTeam:Carnegie Mellon - Revision history2024-03-28T12:43:01ZRevision history for this page on the wikiMediaWiki 1.16.0http://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=297495&oldid=prevYchoo at 03:26, 27 October 20122012-10-27T03:26:47Z<p></p>
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</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=297101&oldid=prevYchoo at 03:13, 27 October 20122012-10-27T03:13:10Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><strong>Figure 1: Measured transcription (left panel) and translation (right panel) rate constants of three new promoters using a new fluorogen-activated biosensor. </strong></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><strong>Figure 1: Measured transcription (left panel) and translation (right panel) rate constants of three new promoters using a new fluorogen-activated biosensor. </strong></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><br> Based on established parts, we have developed a new biosensor that can report levels of both RNA and protein in a single cell. This biosensor enables non-invasive and real-time measurements of RNA and protein expression rates. We have applied the biosensor in the characterization of three new T7Lac promoters, which yielded high quality time-lapse data of both RNA and protein levels (see details in Methods & Results). The data was used to estimate transcription and translation rate constants (see details in Modeling). </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><br> Based on established parts, we have developed a new biosensor that can report levels of both RNA and protein in a single cell. This biosensor enables non-invasive and real-time measurements of RNA and protein expression rates. We have applied the biosensor in the characterization of three new T7Lac promoters, which yielded high quality time-lapse data of both RNA and protein levels (see details in <ins class="diffchange diffchange-inline"><a href = "https://2012.igem.org/Team:Carnegie_Mellon/Met-Overview"> </ins>Methods & Results <ins class="diffchange diffchange-inline"></a></ins>). The data was used to estimate transcription and translation rate constants (see details in <ins class="diffchange diffchange-inline"><a href =" https://2012.igem.org/Team:Carnegie_Mellon/Mod-Overview"> </ins>Modeling <ins class="diffchange diffchange-inline"></a></ins>). </div></td></tr>
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</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=262391&oldid=prevYchoo at 00:21, 4 October 20122012-10-04T00:21:01Z<p></p>
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</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=261838&oldid=prevYchoo at 23:48, 3 October 20122012-10-03T23:48:43Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li><p> We seek to develop a system that will allow researchers in the field of synthetic biology to accurately measure a variety of metrics in gene expression networks including translational efficiency and transcriptional strength.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><li><p> We seek to develop a system that will allow researchers in the field of synthetic biology to accurately measure a variety of metrics in gene expression networks including translational efficiency and transcriptional strength.</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><li><p> We hypothesize that we can use Spinach (a <del class="diffchange diffchange-inline">fluorescent </del>RNA sequence) and a FAP (fluorogen activating protein) as biosensors to <del class="diffchange diffchange-inline">reflect </del>these metrics <i>in vivo</i> (in living cells), rather than <i>in vitro</i> (in a test tube), which can be very costly and <del class="diffchange diffchange-inline">impractical</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><li><p> We hypothesize that we can use Spinach (a <ins class="diffchange diffchange-inline">fluorogen-activating </ins>RNA sequence) and a FAP (fluorogen activating protein) as biosensors to <ins class="diffchange diffchange-inline">measure </ins>these <ins class="diffchange diffchange-inline">gene expression </ins>metrics <i>in vivo</i> (in living cells), rather than <i>in vitro</i> (in a test tube), which can be very costly and <ins class="diffchange diffchange-inline">labor intensive</ins>.</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><li><p> We <del class="diffchange diffchange-inline">will </del>characterize the relationship between <del class="diffchange diffchange-inline">the </del>rates <del class="diffchange diffchange-inline">of production </del>of Spinach and <del class="diffchange diffchange-inline">FAP </del>and the <del class="diffchange diffchange-inline">gene's translational efficiency </del>and <del class="diffchange diffchange-inline">transcription rate</del>. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><li><p> We <ins class="diffchange diffchange-inline">aim to </ins>characterize the relationship between <ins class="diffchange diffchange-inline">synthesis </ins>rates of Spinach and <ins class="diffchange diffchange-inline">transcription rates </ins>and the <ins class="diffchange diffchange-inline">relationship between synthesis rates of FAP </ins>and <ins class="diffchange diffchange-inline">translation rates</ins>. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3><b>Experimental</b></h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><h3><b>Experimental</b></h3></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>The design and implementation of synthetic biological systems often require information on transcription and translation rates and on the impact of both RNA and protein levels on metabolic activities of host cells. Such information is needed when both strong and low levels of expression are desired, depending on the biologists’ goal, e.g. high production or <del class="diffchange diffchange-inline">cell </del>localization of a protein, respectively. To date, however, quantitative information about the expression strength of a promoter is difficult to obtain due to the lack of noninvasive and quick approaches to measure <del class="diffchange diffchange-inline">the </del>levels of RNA and protein in cells. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>The design and implementation of synthetic biological systems often require information on transcription and translation rates and on the impact of both RNA and protein levels on metabolic activities of host cells. Such information is needed when both strong and low levels of expression are desired, depending on the biologists’ goal, e.g.<ins class="diffchange diffchange-inline">, </ins>high production or <ins class="diffchange diffchange-inline">single-molecule </ins>localization of a protein, respectively. To date, however, quantitative information about the expression strength of a promoter is difficult to obtain due to the lack of noninvasive and quick approaches to measure levels of RNA and protein in cells. </div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Here, we engineer a fluorescence-based <del class="diffchange diffchange-inline">sensor </del>that can provide information on both transcription strength and translation efficiency that is noninvasive, easily applied to a variety of promoters, and capable of providing results in a time frame that is short when compared to current technologies. The sensor is based on the use of an RNA aptamer (termed Spinach) and a fluorogen activating protein (FAP). Both the Spinach and FAP become fluorescent in response to binding with dye molecules. The combination of FAP and Spinach will allow us to quantitatively determine relationships involving mRNA and protein, such as translational efficiency.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Here, we engineer a fluorescence-based <ins class="diffchange diffchange-inline">biosensor </ins>that can provide information on both transcription strength and translation efficiency that is noninvasive, easily applied to a variety of promoters, and capable of providing results in a time frame that is short when compared to current technologies. The sensor is based on the use of an RNA aptamer (termed Spinach) and a fluorogen activating protein (FAP). Both the Spinach and FAP become fluorescent in response to binding with dye molecules. The combination of FAP and Spinach will allow us to quantitatively determine relationships involving mRNA and protein, such as translational efficiency.</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>To demonstrate the utility of the sensor, we <del class="diffchange diffchange-inline">will construct </del>and <del class="diffchange diffchange-inline">characterize several T7/Lac </del>promoters. <del class="diffchange diffchange-inline"> </del>For each of the promoters, we <del class="diffchange diffchange-inline">will measure the </del>mRNA and protein fluorescence <del class="diffchange diffchange-inline">during synthesis and after the synthesis ceased as a function of the concentration of dyes added to the cells</del>. The time <del class="diffchange diffchange-inline">dependent </del>fluorescence <del class="diffchange diffchange-inline">measurements </del>of mRNA and protein <del class="diffchange diffchange-inline">levels will be </del>used in a model <del class="diffchange diffchange-inline">that allows one to calculate two important characteristics </del>of <del class="diffchange diffchange-inline">gene expression, namely the polymerase per second (PoPS) </del>and <del class="diffchange diffchange-inline">translational efficiency. Information about other characteristics of the cell, such as degradation </del>constants <del class="diffchange diffchange-inline">for mRNA and protein, and transcriptional efficiency, will be obtained indirectly</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>To demonstrate the utility of the sensor, we <ins class="diffchange diffchange-inline">have constructed </ins>and <ins class="diffchange diffchange-inline">characterized four T7Lac </ins>promoters. For each of the promoters, we <ins class="diffchange diffchange-inline">have measured both </ins>mRNA and protein fluorescence <ins class="diffchange diffchange-inline">over time</ins>. The time<ins class="diffchange diffchange-inline">-lapse </ins>fluorescence <ins class="diffchange diffchange-inline">levels </ins>of mRNA and protein <ins class="diffchange diffchange-inline">were </ins>used in a <ins class="diffchange diffchange-inline">mathematical </ins>model <ins class="diffchange diffchange-inline">for the estimation </ins>of <ins class="diffchange diffchange-inline">transcription </ins>and <ins class="diffchange diffchange-inline">translation rate </ins>constants.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline"></p></del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">We have submitted these </ins>promoters <ins class="diffchange diffchange-inline">to the parts registry</ins>, whose strength is measured by the newly developed <ins class="diffchange diffchange-inline">biosensor</ins>.</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">The outcome of this project will consist of a family of </del>promoters, whose strength is measured by the newly developed <del class="diffchange diffchange-inline">sensor, which covers a relatively broad range of strengths</del>. <del class="diffchange diffchange-inline"> </del></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><i><a href="https://2012.igem.org/Team:Carnegie_Mellon/Met-Overview"> Learn more here</a></i></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><i><a href="https://2012.igem.org/Team:Carnegie_Mellon/Met-Overview"> Learn more here</a></i></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><p>The promoters we submit <del class="diffchange diffchange-inline">will be </del>characterized with these properties. </p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline"><br></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><p>The promoters we submit <ins class="diffchange diffchange-inline">were </ins>characterized with these properties. </p></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><strong>Figure 1: Measured transcription (left panel) and translation (right panel) rate constants of three new promoters using a new fluorogen-activated biosensor. </strong> Based on established parts, we have developed a new biosensor that can report levels of both RNA and protein in a single cell. This biosensor enables non-invasive and real-time measurements of RNA and protein expression rates. We have applied the biosensor in the characterization of three new T7Lac promoters, which yielded high quality time-lapse data of both RNA and protein levels (see details in Methods & Results). The data was used to estimate transcription and translation rate constants (see details in Modeling). </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><strong>Figure 1: Measured transcription (left panel) and translation (right panel) rate constants of three new promoters using a new fluorogen-activated biosensor. </strong></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"><br> </ins>Based on established parts, we have developed a new biosensor that can report levels of both RNA and protein in a single cell. This biosensor enables non-invasive and real-time measurements of RNA and protein expression rates. We have applied the biosensor in the characterization of three new T7Lac promoters, which yielded high quality time-lapse data of both RNA and protein levels (see details in Methods & Results). The data was used to estimate transcription and translation rate constants (see details in Modeling). </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><p> As part of our project, we seek to intrigue high school students about synthetic biology and engineering. In this pursuit, we developed an electrical analog of our BioBricks (with a simulated microscope using LEDs and a photoresistor) to teach high school students about:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><p> As part of our project, we seek to intrigue high school students about synthetic biology and engineering. In this pursuit, we developed an electrical analog of our BioBricks (with a simulated microscope using LEDs and a photoresistor) to teach high school students about:</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><ol><li> Synthetic biology and its relationship to <del class="diffchange diffchange-inline">Biology </del>and <del class="diffchange diffchange-inline">Science and Engineering </del>in general</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ol><li> Synthetic biology and its relationship to <ins class="diffchange diffchange-inline">biology, science, </ins>and <ins class="diffchange diffchange-inline">engineering </ins>in general</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div></li><li> Gene expression and the central dogma of molecular biology</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div></li><li> Gene expression and the central dogma of molecular biology</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div></li><li> How synthetic biologists tackle real-world problems</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div></li><li> How synthetic biologists tackle real-world problems</div></td></tr>
</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=261723&oldid=prevYchoo at 23:39, 3 October 20122012-10-03T23:39:57Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Human Practices</a></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Human Practices</a></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Overview</a></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Overview</a></div></td></tr>
</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=261718&oldid=prevYchoo at 23:39, 3 October 20122012-10-03T23:39:48Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Overview</a></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Overview</a></div></td></tr>
</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=261716&oldid=prevYchoo at 23:39, 3 October 20122012-10-03T23:39:40Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Overview</a></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Overview">Overview</a></div></td></tr>
</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=261712&oldid=prevYchoo at 23:39, 3 October 20122012-10-03T23:39:26Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Software">Software</a></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Software">Software</a></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;"> <a href="https://2012.igem.org/Team:Carnegie_Mellon/Hum-Teach">Teaching Presentation</a></ins></div></td></tr>
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</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=259038&oldid=prevYchoo at 18:43, 3 October 20122012-10-03T18:43:09Z<p></p>
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</table>Ychoohttp://2012.igem.org/wiki/index.php?title=Team:Carnegie_Mellon&diff=259018&oldid=prevYchoo at 18:40, 3 October 20122012-10-03T18:40:21Z<p></p>
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</table>Ychoo