Biomod/2013/Aarhus/Achievements And Future Work: Difference between revisions
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The photosensitizer was successfully synthesized in five steps from the extracted natural compound and subsequently conjugated to DNA. A cholesterol derivative with an NHS-ester handle was synthetized and subsequently coupled to amine modified DNA strands. Furthermore, to achieve a more direct way of labeling DNA with cholesterols, work will be continued in optimizing the triphosphate synthesis and the following purification. This can be used to synthesize a cholesterol modified nucleoside triphosphate which can be used in a cholesterol labeling-kit with TdT. | The photosensitizer was successfully synthesized in five steps from the extracted natural compound and subsequently conjugated to DNA. A cholesterol derivative with an NHS-ester handle was synthetized and subsequently coupled to amine modified DNA strands. Furthermore, to achieve a more direct way of labeling DNA with cholesterols, work will be continued in optimizing the triphosphate synthesis and the following purification. This can be used to synthesize a cholesterol modified nucleoside triphosphate which can be used in a cholesterol labeling-kit with TdT. | ||
===sisiRNA | ===sisiRNA=== | ||
PEG polymers of two different sizes | PEG polymers of two different sizes were conjugated to a sisiRNA strand. | ||
===System in action=== | ===System in action=== |
Revision as of 11:42, 26 May 2015
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Achievements and future work
Origami
Two DNA origamis were designed: a two-layered base plate and a dome that could function as a lid. These were successfully folded and characterized using gel electrophoresis, AFM and TEM. However, connecting the two origamis was not achieved within the time frame of the BIOMOD competition.
Following the BIOMOD competition more experiments in connecting the plate to the dome will be performed, both using staple strands and the peptide lock. Furthermore, the complete structure with its peptide locks will need testing, to see if it can open in response to cleavage by MMP2, possibly by use of Förster resonance energy transfer (FRET).
Peptide lock
The synthesis of the peptide lock was attempted using a gradual buildup of DNA, but was unsuccesful. Coupling of one DNA strand of the lock to the peptide was achieved using click reactions, but the amide coupling of the amine-modified second DNA strand to the peptide was not achieved. After the BIOMOD competition, amide coupling with standard peptide reagents (HBTU, HOBt, etc.), have been planned to take place in order to obtain the desired molecule. Parallel a design using the succesfull click reaction will be investigated.
Chemical modifications
The photosensitizer was successfully synthesized in five steps from the extracted natural compound and subsequently conjugated to DNA. A cholesterol derivative with an NHS-ester handle was synthetized and subsequently coupled to amine modified DNA strands. Furthermore, to achieve a more direct way of labeling DNA with cholesterols, work will be continued in optimizing the triphosphate synthesis and the following purification. This can be used to synthesize a cholesterol modified nucleoside triphosphate which can be used in a cholesterol labeling-kit with TdT.
sisiRNA
PEG polymers of two different sizes were conjugated to a sisiRNA strand.
System in action
The ability of the photosensitizers to induce apoptosis in cells was investigated and it was shown that photosensitizer-DNA alone could not kill cells, but was functional in complex with a cholesterol-DNA. Future experiments will include expanding the cell analyses to anneal the photosensitizer-DNA conjugate into the origami plate and investigate the ability of this system to induce apoptosis. Cholesterol- and photosensitizer modified DNA strands were successfully incorporated into an origami plate. Cells were treated with the cholesterol-modified plates and visualized using confocal microscopy. As no origamis were visible in these images due to low concentrations of the sample, this experiment will be repeated. For the plate with attached sisiRNA-CPP conjugates, annealing of the constructs onto the plate was achieved with a doubly CPP-modified sisiRNA through staple strand overhangs. The plate was tested on cells, both with and without a transfection agent. It was not concluded if the CPPs enabled a cellular uptake, as the sisiRNA could not be liberated outside the cells without a functional peptide lock. However, the disulfide bridges in the sisiRNA design appeared to function as intended, and liberate the sisiRNA-CPP conjugates inside the cell, where it was shown to be functional.
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</style> </head> <body> <div id="indexing"> <div id="sitemap"> <p id="sitemapTitle">SITEMAP | BIOMOD 2013 NANO CREATORS | Aarhus University</p> <div id="footer-contents"> <div class="footer-section"> <p class="footer-section-title">INTRODUCTION</p> <ul> <li><a href="/wiki/Biomod/2013/Aarhus">Home, abstract, animation and video</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Introduction">Introduction</a></li </ul> </div> <div class="footer-section"> <p class="footer-section-title">RESULTS AND DISCUSSION</p> <ul> <li><a href="/wiki/Biomod/2013/Aarhus/Results_And_Discussion/Origami">Origami</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Results_And_Discussion/Peptide_lock">Peptide lock</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Results_And_Discussion/Chemical_Modification">Chemical modification</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Results_And_Discussion/sisiRNA">sisiRNA</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Results_And_Discussion/System_In_Action">System in action</a></li> </ul> </div> <div class="footer-section"> <p class="footer-section-title">MATERIALS AND METHODS</p> <ul> <li><a href="/wiki/Biomod/2013/Aarhus/Materials_And_Methods/Origami">Origami</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Materials_And_Methods/Peptide_lock">Peptide lock</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Materials_And_Methods/Chemical_Modification">Chemical modification</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Materials_And_Methods/sisiRNA">sisiRNA</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Materials_And_Methods/System_In_Action">System in action</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Materials_And_Methods/Methods">Methods</a></li> </ul> </div> <div class="footer-section"> <p class="footer-section-title">SUPPLEMENTARY</p> <ul> <li><a href="/wiki/Biomod/2013/Aarhus/Supplementary/Team_And_Acknowledgments">Team and acknowledgments</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Supplementary/Optimizations">Optimizations</a></li> <li><a href="/wiki/Biomod/2013/Aarhus/Supplementary/Supplementary_Data">Supplementary data</a></li>
<li><a
href="/wiki/Biomod/2013/Aarhus/Supplementary/Supplementary_Informations">Supplementary informations</a> <li><a href="/wiki/Biomod/2013/Aarhus/Supplementary/References">References</a></li> </ul> </div> </div> <div> <p id="copyright">Copyright (C) 2013 | BIOMOD Team Nano Creators @ Aarhus University | Programming by: <a href="mailto:pvskaarup@gmail.com?Subject=BIOMOD 2013:">Peter Vium Skaarup</a>.</p> </div> </div>
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