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− | <tr class="lastRow"><td align=center valign=center colspan=3> | + | <tr class="lastRow"><td align=center valign=center colspan=6> |
− | <h3>First semester welcome program at Weihenstephan, TU Munich (10th October 2017)</h3> | + | <h3>Marienplatz public interaction (3rd September 2017)</h3> |
| <p> | | <p> |
− | As human practices team, we informed the freshmen, who study in major molecular biotechnology and biology at Technical University of Munich, about iGEM competition and synthetic biology because we think that they should be interested in iGEM competition. So we would like to give them an opportunity to start to do something in this field because it is better to get the information early.There were around 200 students present and we were able to give them some insights to the iGEM world.</p>
| + | We believe that the best way of making people aware of synthetic biology and its future perspective is public interaction. Most ethical issues can be overcome by informing the public about the subject. For that sake, we went to the center of Munich, Marienplatz which is famous not only for the locals but also it is a hub for tourists from all over the world. We took our team members along with our camera and survey sheets to ask the public about their view on synthetic biology and modified organisms. We also explained them about our project design and asked them if they would be interested if we had such a device in the market. The people seem to be very excited about it and were willing to buy such simple lab-on-a-chip devices if they become available in the future. |
− | </td>
| + | |
− | <td align=center valign=center colspan=3>
| + | |
− | <h3>First semester welcome program at Martinesried, LMU (11th October 2017)</h3>
| + | |
− | <p>
| + | |
− | Two people of the team gave a presentation about iGEM and the project we were working on. Over 140 first semester students from the LMU and TU Munich came there and listened to them. Most of the students were very motivated afterwards to join the iGEM competition in the following years, because it has a lot to offer and its scientific fields in synthetic biology are nearly infinite. With them we discussed about the future of synthetic biology and also got great positive feedback for our idea. So we hope that some of them will enrich the future of the Munich teams.</p>
| + | |
− | </td>
| + | |
− | </tr>
| + | |
− | | + | |
− | | + | |
− | <tr><td colspan=6 align=center valign=center>
| + | |
− | <h3>Colorimetric read-outs</h3>
| + | |
− | <p>
| + | |
− | To couple CascAID with an easy read-out method we explored three colorimetric read-outs:
| + | |
| </p> | | </p> |
| </td> | | </td> |
| </tr> | | </tr> |
| | | |
− | <tr><td colspan=2 align=center valign=center> | + | <tr class="lastRow"><td align=center valign=center colspan=6> |
| + | <h3>Nano day (10th September 2017)</h3> |
| <p> | | <p> |
− | <b>AeBlue</b>: The RNA strand in a specially designed RNA/DNA dimer is cut by Cas13a's collateral
| + | The Nano day is a special program organized annually by NIM (Nanosystems Initiative Munich). Students from different labs working in physics, biophysics, and synthetic biology were present with their posters. The participation of interested people was really high. There were also do it yourself session and displays of different technologies so that the general people and students could try it themselves. A general quiz for everyone was also part of the program and at the end the winners of the quiz were awarded with gifts. There were also talks from renowned professors and researchers on different topics. |
− | activity. After digestion, the interaction between the two strands is too weak to hold the dimer and it
| + | People showed great interest in our work and were willing to participate in our survey as well. The program took place from 11:00 am to 18:00 pm and was open and free for everyone. |
− | decays. We can then use the DNA-strand as template to translate the chromoprotein <a href="http://parts.igem.org/Part:BBa_K864401">aeBlue</a>.
| + | |
− | </p>
| + | |
− | </td>
| + | |
− | <td colspan=4 align=center valign=center>
| + | |
− | <img src="https://static.igem.org/mediawiki/2017/9/90/T--Munich--Description_aeBlue.svg">
| + | |
− | <p>Diagram of aeBlue</p>
| + | |
− | </td>
| + | |
− | </tr>
| + | |
− | | + | |
− | <tr><td colspan=2 align=center valign=center>
| + | |
− | <p>
| + | |
− | <b>Intein-Extein</b>: By binding TEV-protease with a RNA-linker we can use Cas13a's collateral activity
| + | |
− | to regulate the protease's diffusion and use it to cleave a TEV tag separating the intein regions of a
| + | |
− | modified chromophore. After the first cleavage, the intein segment excises itself<sup><a class="myLink" href="#13">13</a></sup>, bringing together the
| + | |
− | halves of the chromophore. Only then is the chromophore functional and produces the colorimetric
| + | |
− | read-out.
| + | |
− | </p>
| + | |
− | </td>
| + | |
− | <td colspan=4 align=center valign=center>
| + | |
− | <a href="http://www.uni-muenchen.de/studium/lehre_at_lmu/index.html"><img src="https://static.igem.org/mediawiki/2017/9/9a/T--Munich--Logo_LehreLMU.gif" width="200"></a>
| + | |
− | <p>Diagram of Intein-Extein</p>
| + | |
− | </td>
| + | |
− | </tr>
| + | |
− | | + | |
− | <tr class="lastRow"><td colspan=2 align=center valign=center>
| + | |
− | <p>
| + | |
− | <b>Gold nanoparticles</b>: Gold nanoparticles coated with short DNA sequences are held closely
| + | |
− | together by a complementary linker RNA, which makes the solution intense blue<sup><a class="myLink" href="#14">14</a></sup>. Activated Cas13a cuts
| + | |
− | the linker RNA, causing the nanoparticles to diffuse away from each other. This increase in distance
| + | |
− | causes a color change to intense red.
| + | |
− | </p>
| + | |
− | </td>
| + | |
− | <td colspan=4 align=center valign=center>
| + | |
− | <img src="https://static.igem.org/mediawiki/2017/b/b3/T--Munich--Description_Goldnanoparticles.svg">
| + | |
− | <p>Gold nanoparticles</p>
| + | |
− | </td>
| + | |
− | </tr>
| + | |
− | | + | |
− | <tr><td colspan=6 align=center valign=center>
| + | |
− | <h3>Software</h3>
| + | |
− | <p>
| + | |
− | To help facilitate the design of crRNA, the sequences that give CascAID its specificity, we developed a
| + | |
− | software tool that checks crRNA for unwanted secondary structures. This gives valuable insight on
| + | |
− | whether the sequence is suited to use with Cas13a or whether some modifications are needed.
| + | |
− | Together with Team Delft's software tool which designs the corresponding crRNA based on the target,
| + | |
− | we collaborated to develop a powerful tool that suggests crRNA sequences and checks their usability
| + | |
| </p> | | </p> |
| </td> | | </td> |
| </tr> | | </tr> |
− |
| |
− | <tr><td colspan=6 align=center valign=center>
| |
− | <h3>References</h3>
| |
− | <p>
| |
− | <ol style="text-align: left">
| |
− | <li id="ref_1">Cohen, Limor, and David R. Walt. "Single-Molecule Arrays for Protein and Nucleic Acid Analysis." Annual Review of Analytical Chemistry 0 (2017).</li>
| |
− | <li id="ref_2">Nakano, Michihiko, et al. "Single-molecule PCR using water-in-oil emulsion." Journal of biotechnology 102.2 (2003): 117-124.</li>
| |
− | <li id="ref_3">Taniguchi, Yuichi, et al. "Quantifying E. coli proteome and transcriptome with single-molecule sensitivity in single cells." science 329.5991 (2010): 533-538.</li>
| |
− | <li id="ref_4">Rissin, David M., et al. "Single-molecule enzyme-linked immunosorbent assay detects serum proteins at subfemtomolar concentrations." Nature biotechnology 28.6 (2010): 595-599.</li>
| |
− | <li id="ref_5">Pardee, Keith, et al. "Rapid, low-cost detection of Zika virus using programmable biomolecular components." Cell 165.5 (2016): 1255-1266.</li>
| |
− | <li id="ref_6">Slomovic, Shimyn, Keith Pardee, and James J. Collins. "Synthetic biology devices for in vitro and in vivo diagnostics." Proceedings of the National Academy of Sciences 112.47 (2015): 14429-14435.</li>
| |
− | <li id="ref_7">Tang, Ruihua, et al. "A fully disposable and integrated paper-based device for nucleic acid extraction, amplification and detection." Lab on a Chip 17.7 (2017): 1270-1279.</li>
| |
− | <li id="ref_8">Vashist, Sandeep Kumar, et al. "Emerging technologies for next-generation point-of-care testing." Trends in biotechnology 33.11 (2015): 692-705.</li>
| |
− | <li id="ref_9">Gubala, Vladimir, et al. "Point of care diagnostics: status and future." Analytical chemistry 84.2 (2011): 487-515.</li>
| |
− | <li id="ref_10">Abudayyeh, Omar O., et al. "C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector." Science 353.6299 (2016): aaf5573.</li>
| |
− | <li id="ref_11">Gootenberg, Jonathan S., et al. "Nucleic acid detection with CRISPR-Cas13a/C2c2." Science (2017): eaam9321.</li>
| |
− | <li id="ref_12">https://www.idtdna.com/pages/docs/technical-reports/in_vitro_nuclease_detectionD325FDB69855.pdf (retrieved: 13.10.17)</li>
| |
− | <li id="ref_13"> Anraku, Yasuhiro, Ryuta Mizutani, and Yoshinori Satow. "Protein splicing: its discovery and structural insight into novel chemical mechanisms." IUBMB life 57.8 (2005): 563-574.</li>
| |
− | <li id="ref_14">Link, Stephan, and Mostafa A. El-Sayed. "Size and temperature dependence of the plasmon absorption of colloidal gold nanoparticles." The Journal of Physical Chemistry B 103.21 (1999): 4212-4217.</li>
| |
− | </ol>
| |
− | </p>
| |
− | </td>
| |
− | </tr>
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