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<li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Team" onmouseover="mopen('m2')" onmouseout="mclosetime()" style="padding:15px 0px;">Team</a>
<li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Team" onmouseover="mopen('m2')" onmouseout="mclosetime()" style="padding:15px 0px;">Team</a>
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<li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs" onmouseover="mopen('m3')" onmouseout="mclosetime()" style="padding:15px 0px;">Designs</a>
<li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs" onmouseover="mopen('m3')" onmouseout="mclosetime()" style="padding:15px 0px;">Designs</a>
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                <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs">Background</a>
<a href="#">Item2</a>
<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/Self-assemblyTrack">Self-assembly Track</a>
<a href="#">Item3</a>
<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/DeliveryDevice">The Delivery Device</a>
<a href="#">Item4</a>
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<li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results" onmouseover="mopen('m4')" onmouseout="mclosetime()" style="height:35px;padding:8px 0px 7px 0px;">Experiments &amp; Results</a>
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<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results">Polymerizing</a>
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<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results/Cleavage">Cleavage</a>
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<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results/DeliveryDevice">Delivery Device</a>
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     <li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Attributions" onmouseover="mopen('m6')" onmouseout="mclosetime()" style="padding:15px 0px;">Attributions</a>
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<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs" style="font-size:14px;"><b>Background</b></a>
<p>HCR、DNAzyme、DNA-walker、Origami</p>
<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/Self-assemblyTrack" style="font-size:14px;"><b>Self-assembly Track</b></a>
<p>T1-T4、The polymerization reaction</p>
<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/DeliveryDevice"><b>The Delivery device</b></a>
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<span style="color:#000;font-size:22px;"><b>Background</b></span>
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<table id="toc" class="toc" summary="Contents" style="font-family:Calibri;float:right;font-size:120%;"><tbody><tr><td><div id="toctitle"><h2>Contents</h2></div>
<table id="toc" class="toc" summary="Contents" style="font-family:Calibri;float:right;font-size:120%;"><tbody><tr><td><div id="toctitle"><h2>Contents</h2></div>
<ul>
<ul>
<li class="toclevel-1"><a href="#The_DNA.E2.80.94Track"><span class="tocnumber">1</span> <span class="toctext">The DNA—Track</span></a></li>
<li class="toclevel-1"><a href="#HCR"><span class="tocnumber">1</span> <span class="toctext">HCR</span></a></li>
<li class="toclevel-1"><a href="#The_Walker"><span class="tocnumber">2</span> <span class="toctext">The Walker</span></a></li>
<li class="toclevel-1"><a href="#DNAzyme"><span class="tocnumber">2</span> <span class="toctext">DNAzyme</span></a></li>
<li class="toclevel-1"><a href="#Agarose_electrophoresis"><span class="tocnumber">3</span> <span class="toctext">Agarose electrophoresis</span></a></li>
<li class="toclevel-1"><a href="#DNA-walker"><span class="tocnumber">3</span> <span class="toctext">DNA-walker</span></a></li>
<li class="toclevel-1"><a href="#PAGE"><span class="tocnumber">4</span> <span class="toctext">PAGE</span></a>
<li class="toclevel-1"><a href="#Origami"><span class="tocnumber">4</span> <span class="toctext">Origami</span></a>
<ul>
<li class="toclevel-2"><a href="#Materials"><span class="tocnumber">4.1</span> <span class="toctext">Materials</span></a></li>
<li class="toclevel-2"><a href="#Methods"><span class="tocnumber">4.2</span> <span class="toctext">Methods</span></a></li>
</ul>
</li>
<li class="toclevel-1"><a href="#PCR_Annealing_Procedure_.28for_Logic_Gate.29"><span class="tocnumber">5</span> <span class="toctext">PCR Annealing Procedure (for Logic Gate)</span></a></li>
<li class="toclevel-1"><a href="#Recipe"><span class="tocnumber">6</span> <span class="toctext">Recipe</span></a></li>
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= HCR =
= HCR =


HCR: Stable DNA monomers assemble only upon exposure to atarget DNA fragment. In the simplest version of this process, two stable species of DNA hairpins coexist in solution until the introduction of initiator strands triggers a cascade of hybridization events that yields nicked double helices analogous to alternating copolymers. The average molecular weight of the HCR products varies inversely with initiator concentration. Amplification of more diverse recognition events can be achieved by coupling HCR to aptamer triggers. This functionality allows DNA to act as an amplifying transducer for biosensing applications.
Stable DNA monomers assemble only upon exposure to atarget DNA fragment. In the simplest version of this process, two stable species of DNA hairpins coexist in solution until the introduction of initiator strands triggers a cascade of hybridization events that yields nicked double helices analogous to alternating copolymers. The average molecular weight of the HCR products varies inversely with initiator concentration. Amplification of more diverse recognition events can be achieved by coupling HCR to aptamer triggers. This functionality allows DNA to act as an amplifying transducer for biosensing applications.


[[Image:#.png |thumb|400px| '''Figure2.1.1 (a–c)''' Secondary structure schematic of HCR function. Letters marked with * are complementary to the corresponding unmarked letter.(a) Hairpins H1 and H2 are stable in the absence of initiator I. (b) I nucleates at the sticky end of H1 and undergoes an unbiased strand displacement interaction to open the hairpin. (c) The newly exposed sticky end of H1 nucleates at the sticky end of H2 and opens the hairpin to expose a sticky end on H2 that is identical in sequence to I. Hence, each copy of I can propagate a chain reaction of hybridization events between alternating H1 and H2 hairpins to form a nicked double-helix, amplifying the signal of initiator binding. Figure and introduction from <b>Robert M. Dirks and Niles A. Pierce</b> <sub>PNAS  October 26, 2004  vol. 101 no. 43</sub>]]
[[Image:TJU-figure2.1.1.png|thumb|600px|center|'''Figure2.1.1 (a–c)''' Secondary structure schematic of HCR function. Letters marked with * are complementary to the corresponding unmarked letter.(a) Hairpins H1 and H2 are stable in the absence of initiator I. (b) I nucleates at the sticky end of H1 and undergoes an unbiased strand displacement interaction to open the hairpin. (c) The newly exposed sticky end of H1 nucleates at the sticky end of H2 and opens the hairpin to expose a sticky end on H2 that is identical in sequence to I. Hence, each copy of I can propagate a chain reaction of hybridization events between alternating H1 and H2 hairpins to form a nicked double-helix, amplifying the signal of initiator binding. Figure and introduction from <b>Robert M. Dirks and Niles A. Pierce</b> <sub>PNAS  October 26, 2004  vol. 101 no. 43</sub>]]


= DNAzyme =
= DNAzyme =
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DNAzymes (also known as deoxyribozymes, DNA enzymes or catalytic DNA, are DNA molecules that have the ability to perform a chemical reaction, such as catalytic action. Since the description of the first DNAzyme for the cleavage of RNA in 1994, many more DNAzymes have been reported to catalyze many different types of chemical transformations, such as porphyrin metalation, DNA phosphorylation, RNA ligation, thymine-thymine dimer repair, carbon-carbon bond formation, and hydrolytic cleavage of DNA. DNA is chemically stable and can be conveniently produced by highly efficient automated DNA synthesis. Therefore, DNAzymes can be quite useful in research and applications in chemical biology, biotechnology, and medical areas.
DNAzymes (also known as deoxyribozymes, DNA enzymes or catalytic DNA, are DNA molecules that have the ability to perform a chemical reaction, such as catalytic action. Since the description of the first DNAzyme for the cleavage of RNA in 1994, many more DNAzymes have been reported to catalyze many different types of chemical transformations, such as porphyrin metalation, DNA phosphorylation, RNA ligation, thymine-thymine dimer repair, carbon-carbon bond formation, and hydrolytic cleavage of DNA. DNA is chemically stable and can be conveniently produced by highly efficient automated DNA synthesis. Therefore, DNAzymes can be quite useful in research and applications in chemical biology, biotechnology, and medical areas.


[[Image:#.png|thumb|400px| '''Figure 2.1.2'''The sequence of 8-17 DNAzyme.The triangle represents the cutting site. (From Peracchi A <i>et al., J. Mol. Biol.</i>, 2005(352): 783–794.)]]
[[Image:TJU-figure2.1.2.png|thumb|center|400px| '''Figure 2.1.2''' The sequence of 8-17 DNAzyme.The triangle represents the cutting site. (From Peracchi A <i>et al., J. Mol. Biol.</i>, 2005(352): 783–794.)]]


The Cu<sup>2+</sup> DNAzyme is also an ssDNA that contains a stem-loop of 8 base-pairing. The catalytic domain consists of a conservative sequence of six basepair. The two binding arms flanking the catalytic domain bind with the substrate, one of which forms a DNA triplex of the stem-loop with the substrate. Unlike 8-17, the substrate of Cu<sup>2+</sup> DNAzyme is deoxyribonucleotide. When the Cu<sup>2+</sup>concentration is <1μM, DNAzyme is still activated. When other ions’ concentration is enormously bigger than Cu<sup>2+</sup>, the DNAzyme still didn’t recover its full activity, which shows its great selectivity of Cu<sup>2+</sup>.
The Cu<sup>2+</sup> DNAzyme is also an ssDNA that contains a stem-loop of 8 base-pairing. The catalytic domain consists of a conservative sequence of six basepair. The two binding arms flanking the catalytic domain bind with the substrate, one of which forms a DNA triplex of the stem-loop with the substrate. Unlike 8-17, the substrate of Cu<sup>2+</sup> DNAzyme is deoxyribonucleotide. When the Cu<sup>2+</sup>concentration is <1μM, DNAzyme is still activated. When other ions’ concentration is enormously bigger than Cu<sup>2+</sup>, the DNAzyme still didn’t recover its full activity, which shows its great selectivity of Cu<sup>2+</sup>.
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The mechanism of the DNA walker with a DNAzyme should be like this.
The mechanism of the DNA walker with a DNAzyme should be like this.


[[Image:#.png|thumb|400px| '''Figure2.1.3'''the mechanism of the walker with DNAzyme]]
[[Image:TJU-figure2.1.3.png|thumb|center|400px| '''Figure2.1.3''' The mechanism of the walker with DNAzyme.]]


=origami=
=Origami=
DNA origami is the nanoscale folding of DNA to create arbitrary two and three dimensional shapes at the nanoscale. The specificity of the interactions between complementary base pairs make DNA a useful construction material, through design of its base sequences.
DNA origami is the nanoscale folding of DNA to create arbitrary two and three dimensional shapes at the nanoscale. The specificity of the interactions between complementary base pairs make DNA a useful construction material, through design of its base sequences.


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This year, we used the design from 2012 Harvard BIOMOD team to build the origami. What’s different is that we load ssDNAs on the staple strand. The ssDNAs can serve as the substrate of logic gate of 8-17 and Cu2+ DNAzyme, thus the release can be controlled by it. This new origami can serve as a miRNA delivery system based on ion detection.
This year, we used the design from 2012 Harvard BIOMOD team to build the origami. What’s different is that we load ssDNAs on the staple strand. The ssDNAs can serve as the substrate of logic gate of 8-17 and Cu2+ DNAzyme, thus the release can be controlled by it. This new origami can serve as a miRNA delivery system based on ion detection.


 
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<br />
<br />
E-mail：austinamens@gmail.com | Address:Building No.20, No.92 Weijin Road, Tianjin, China | Zip-code：300072
E-mail：austinamens@gmail.com | Address:Building No.20, No.92 Weijin Road, Tianjin, China | Zip-code：300072
<br />Copyright 2013 © Tianjin University iGEM Team</p>
<br />Copyright 2013 © Tianjin University Biomod Team</p>
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               <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs">Background</a>

<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/Self-assemblyTrack">Self-assembly Track</a> <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/DeliveryDevice">The Delivery Device</a> </div> </li> <li><a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results" onmouseover="mopen('m4')" onmouseout="mclosetime()" style="height:35px;padding:8px 0px 7px 0px;">Experiments &amp; Results</a> 

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<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results">Polymerizing</a> <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results/Cleavage">Cleavage</a> <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Experiments &amp; Results/DeliveryDevice">Delivery Device</a>

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<a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs" style="font-size:14px;"><b>Background</b></a> <p>HCR、DNAzyme、DNA-walker、Origami</p> <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/Self-assemblyTrack" style="font-size:14px;"><b>Self-assembly Track</b></a> <p>T1-T4、The polymerization reaction</p> <a href="http://openwetware.org/wiki/Biomod/2013/Tianjin/Designs/DeliveryDevice"><b>The Delivery device</b></a> <br/><br/> </div>

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document.getElementById("toc").className="tocbefore"; } else{ document.getElementById("toc").className="tocafter"; } } window.onscroll=getScrollTop; </script> <div style="width:280px;float:left;"> <table id="toc" class="toc" summary="Contents" style="font-family:Calibri;float:right;font-size:120%;"><tbody><tr><td><div id="toctitle"><h2>Contents</h2></div> <ul> <li class="toclevel-1"><a href="#HCR"><span class="tocnumber">1</span> <span class="toctext">HCR</span></a></li> <li class="toclevel-1"><a href="#DNAzyme"><span class="tocnumber">2</span> <span class="toctext">DNAzyme</span></a></li> <li class="toclevel-1"><a href="#DNA-walker"><span class="tocnumber">3</span> <span class="toctext">DNA-walker</span></a></li> <li class="toclevel-1"><a href="#Origami"><span class="tocnumber">4</span> <span class="toctext">Origami</span></a> </ul> </td></tr></tbody></table> </div>


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HCR

Stable DNA monomers assemble only upon exposure to atarget DNA fragment. In the simplest version of this process, two stable species of DNA hairpins coexist in solution until the introduction of initiator strands triggers a cascade of hybridization events that yields nicked double helices analogous to alternating copolymers. The average molecular weight of the HCR products varies inversely with initiator concentration. Amplification of more diverse recognition events can be achieved by coupling HCR to aptamer triggers. This functionality allows DNA to act as an amplifying transducer for biosensing applications.

Figure2.1.1 (a–c) Secondary structure schematic of HCR function. Letters marked with * are complementary to the corresponding unmarked letter.(a) Hairpins H1 and H2 are stable in the absence of initiator I. (b) I nucleates at the sticky end of H1 and undergoes an unbiased strand displacement interaction to open the hairpin. (c) The newly exposed sticky end of H1 nucleates at the sticky end of H2 and opens the hairpin to expose a sticky end on H2 that is identical in sequence to I. Hence, each copy of I can propagate a chain reaction of hybridization events between alternating H1 and H2 hairpins to form a nicked double-helix, amplifying the signal of initiator binding. Figure and introduction from Robert M. Dirks and Niles A. Pierce PNAS October 26, 2004 vol. 101 no. 43

DNAzyme

DNAzymes (also known as deoxyribozymes, DNA enzymes or catalytic DNA, are DNA molecules that have the ability to perform a chemical reaction, such as catalytic action. Since the description of the first DNAzyme for the cleavage of RNA in 1994, many more DNAzymes have been reported to catalyze many different types of chemical transformations, such as porphyrin metalation, DNA phosphorylation, RNA ligation, thymine-thymine dimer repair, carbon-carbon bond formation, and hydrolytic cleavage of DNA. DNA is chemically stable and can be conveniently produced by highly efficient automated DNA synthesis. Therefore, DNAzymes can be quite useful in research and applications in chemical biology, biotechnology, and medical areas.

Figure 2.1.2 The sequence of 8-17 DNAzyme.The triangle represents the cutting site. (From Peracchi A et al., J. Mol. Biol., 2005(352): 783–794.)

The Cu2+ DNAzyme is also an ssDNA that contains a stem-loop of 8 base-pairing. The catalytic domain consists of a conservative sequence of six basepair. The two binding arms flanking the catalytic domain bind with the substrate, one of which forms a DNA triplex of the stem-loop with the substrate. Unlike 8-17, the substrate of Cu2+ DNAzyme is deoxyribonucleotide. When the Cu2+concentration is <1μM, DNAzyme is still activated. When other ions’ concentration is enormously bigger than Cu2+, the DNAzyme still didn’t recover its full activity, which shows its great selectivity of Cu2+.

DNA-walker

DNA walkers are a class of nucleic acid nanomachines that exhibit directional motion along a linear track. A large number of schemes have been demonstrated. One strategy is to control the motion of the walker along the track using control strands that need to be manually added in sequence. Another approach is to make use of restriction enzymes or deoxyribozymes to cleave the strands and cause the walker to move forward, which has the advantage of running autonomously, and we choose this kind of walker this year. A later system could walk upon a two-dimensional surface rather than a linear track, and demonstrated the ability to selectively pick up and move molecular cargo.[55] Additionally, a linear walker has been demonstrated that performs DNA-templated synthesis as the walker advances along the track, allowing autonomous multistep chemical synthesis directed by the walker.

The mechanism of the DNA walker with a DNAzyme should be like this.

Figure2.1.3 The mechanism of the walker with DNAzyme.

Origami

DNA origami is the nanoscale folding of DNA to create arbitrary two and three dimensional shapes at the nanoscale. The specificity of the interactions between complementary base pairs make DNA a useful construction material, through design of its base sequences.

Developed by Paul Rothemund at the California Institute of Technology, the process involves the folding of a long single strand of viral DNA aided by multiple smaller "staple" strands. These shorter strands bind the longer in various places, resulting in various shapes, including a smiley face and a coarse map of China and the Americas, along with many three-dimensional structures such as cubes.

To produce a desired shape, images are drawn with a raster fill of a single long DNA molecule. This design is then fed into a computer program that calculates the placement of individual staple strands. Each staple binds to a specific region of the DNA template, and thus due to Watson-Crick base pairing, the necessary sequences of all staple strands are known and displayed. The DNA is mixed, then heated and cooled. As the DNA cools, the various staples pull the long strand into the desired shape. Designs are directly observable via several methods, including atomic force microscopy, or fluorescence microscopy when DNA is coupled to fluorescent materials.

This year, we used the design from 2012 Harvard BIOMOD team to build the origami. What’s different is that we load ssDNAs on the staple strand. The ssDNAs can serve as the substrate of logic gate of 8-17 and Cu2+ DNAzyme, thus the release can be controlled by it. This new origami can serve as a miRNA delivery system based on ion detection.

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<div class="link"> <p><br /> <br /> E-mail：austinamens@gmail.com | Address:Building No.20, No.92 Weijin Road, Tianjin, China | Zip-code：300072 <br />Copyright 2013 © Tianjin University Biomod Team</p> </div>


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