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<h1>Design</h1>
<h1>Design</h1>
<p>To complete our goal, we need to develop next two systems.</p>
1. Programmable Output System<br>
2. Taste Releasing System<br><br>
<!--
polymerase, nickase, and restriction enzyme.
In addition, to demonstrate these systems, we put taste substances into liposomes and effuse them.</p>
This system aims to output signals to each input signals in order. To achieve this goal,
-->
<h1>1. Programmable Output System </h1>
<p> The purpose of this system is to release output DNA in order.
We propose two different approaches.<br><br>
<h2>1st Approach; Enzyme system</h2>
<p>Enzyme System has three processes.</p>
1. Amplifying Process:The process in which DNA Polymerase amplifies KEY DNA. <br>
2. Releasing Process:The process in which KEY DNA releases the liposome. <br>
3. Renewing Process:The process in which restriction enzyme renews the 3’ end of the input-DNA sequence to get an output. <br>
<img src="http://openwetware.org/images/9/9e/Fde1a0-01.png">
<h3>1.Amplifying process</h3>
<p>
In this process, first, domains A, B, and C in the template combine with domains A, B, and C in the input respectively. Then, polymerase copies 5’ end of the template sequence and extends input sequence. After that, nickase cleaves the end of the copied domain. Then, polymerase works at the gap created by the nickase and push out the domain A. Repeating this process again and again, we amplify the domain A. (Fig.1)
</p>
<p>
This DNA domain, we call it A1, becomes an input to following process 2 and 3.
</p>
<img src="http://openwetware.org/images/c/c9/Figure-01.png"><br><br>
<!--
&#9312; Domains A0, B0, and C0 in the template combine with domains A0-, B0-, and C0- in the input respectively. Input-Template complex is created.<br>
&#9313; Polymerase recognizes 3’ end of input. (Identified with dashed circle) <br>
&#9314; Extending input sequence. The sequence A1 is newly formed. <br>
&#9315; Polymerase runs away from Input-Template complex. <br>
&#9316; Nickase combines with its recognition cite. <br>
&#9317; Cleaving the end of the copied domain./ Making nick between A0 and A1. <br>
&#9318; Polymerase works from the gap (Identified with dashed circle) created by the nickase. <br>
&#9319; Running on DNA with making nucleotides and pushing out the A1. <br>
&#9320; Repeating this process again and again amplifies the number of domain A1. <br><br>
-->
<h3>2.Releasing process</h3>
<p> A1 combines with the 3’ end of the DNA which is combined with the DNA modified with liposome (output-A). Polymerase recognizes the 3’ end of the A1 and then extend the chain.
</p>
<p>
At the same time, output-A is denatured and released.
</p>
<img src="http://openwetware.org/images/1/1f/IMG_3657-02.jpg" width="735px" height="600px">
<p>A1 combines with the end of the DNA which fixes the liposome-modifying DNA (output-A).
Polymerases recognize the 3’end of the A1 and extend the chain.
At the same time, output-A is denatured and released.</p>
<h3>3.Renewing process</h3>
<img src="http://openwetware.org/images/7/7f/Figure3-01.png">
<p> When A1 combines with the gate-A, the DNA which has the recognition sequence corresponding to the restriction enzyme is released by polymerase. When this DNA combines with the structure made of input and template, a restriction enzyme activates, and cleaves the chain between A and B. By doing this, the domain near the 3’ end of the input becomes domain B. Then, the process goes back to the system-1.
</p>
<p> In this way, output-A, output-B, and output-C are released in order.</p>
<h2>2nd Approach; Enzyme-free System</h2>
<img src="http://openwetware.org/images/9/97/Figure_EnzymeFree-01.png">
<img src="http://openwetware.org/images/f/f6/Figure_EnzymeFree-03.png">
<img src="http://openwetware.org/images/8/89/Figure_EnzymeFree-04.png">
<p>This approach is inspired by seesaw gate (Lulu Qian et.al, 2011). Our goal is to get different output signals in order of input signals. In this system , we should arrange a input , a trigger ,fuels, the double strand DNA bonding with liposome.</p>
<img src="http://openwetware.org/images/7/79/Figure_EnzymeFree-02.png">
<p>（用意するもの）
fuel,gate、Liposomeが結合した二本鎖DNA (Fa,b,c Ga,b,c La,b,c)
input(I1),trigger(T1)それぞれ一種類
</p>
<p>Reactions as follows : </p>
<h3>1.</h3>
<p> Input-DNA sequence are dissolved in the solution including trigger , fuels , gates , and the double strand DNA bonding with liposome. Then, 　trigger combines with input and
the single strand DNA (DNA1)which makes input are released. </p>
<h3>2.</h3>
<p> DNA1 combines with the double strand DNA bonding with liposome . In this reaction ,  the single strand DNA bonding with liposome are released due to the difference in length of DNA sequence which makes the double strand DNA.
This reaction causes earlier so that toehold  is longer than that of gate.
</p>
<h3>3.</h3>
<p> Other DNA1 combine with gate(Ga).
the single strand DNA (DNA2)which makes gate(Ga) are released so that the length of the double strand is difference.
In addition , DNA1 combines with gate.
</p>
<h3>4.</h3>
<p> DNA1 is released again so that DNA3 react with fuel(Fa) ,
due to the difference in length of DNA.
</p>
<h3>5.</h3>
<p> When we perform process3 and 4 repeatedly , the number of DNA2 is increasing. DNA2 is the key  of next process.
</p>
<img src="http://openwetware.org/images/4/42/Enzyme-free_System_picture2.jpg" width="441px" height="599px">
<h3>6.</h3>
<p> DNA2 combines with input as trigger , so the reaction said above caused.In this reaction , DNA is  released as DNA2 in the reaction　said above .
</p>
<img src="http://openwetware.org/images/2/20/Enzyme-free_System_picture3.jpg" width="426px" height="600px">
<h3>7.</h3>
<p> DNA4 combines with input as trigger , so the reaction said above caused.
</p>
<img src="http://openwetware.org/images/d/d4/Enzyme-free_System_picture4.jpg" width=395px height="599px">
<p> We can get output signals in order by using this reaction.
</p>
<h2>2. Taste releasing system </h2>
<p>
The purpose of this system is to encapsulate taste substances in liposome and to release liposome attached to substrate.
The DNA which has the complementary sequence (comp DNA) to the Output is sprouted on agarose gel. They are hybridized each other. When A1 reacts with comp DNA, the output is separated from comp DNA, then the liposome is released from the agarose gel, and taste substances are diffused.
</p>
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<body> <div id="globalnav" class="design"> <ul> <li id="gn-home"><a href="/wiki/Biomod/2014/Sendai">Home</a></li> <li id="gn-intro"><a href="/wiki/Biomod/2014/Sendai/Introduction">Introduction</a></li> <li id="gn-design"><a href="/wiki/Biomod/2014/Sendai/Design">Design</a></li> <li id="gn-simu"><a href="/wiki/Biomod/2014/Sendai/Simulation">Simulation</a></li> <li id="gn-xp"><a href="/wiki/Biomod/2014/Sendai/Experiment">Experiment</a></li> <!--<li id="gn-protocol"><a href="/wiki/Biomod/2014/Sendai/Protocol">Protocol</a></li>--> <li id="gn-dis"><a href="/wiki/Biomod/2014/Sendai/Discussion">Discussion</a></li> <li id="gn-team"><a href="/wiki/Biomod/2014/Sendai/Team">Team</a></li> <li id="gn-end"><a href="#"></a></li> </ul> </div> <div id="main"> <h1>Design</h1> <p>To complete our goal, we need to develop next two systems.</p> 1. Programmable Output System<br> 2. Taste Releasing System<br><br>

<!-- polymerase, nickase, and restriction enzyme.

In addition, to demonstrate these systems, we put taste substances into liposomes and effuse them.</p> This system aims to output signals to each input signals in order. To achieve this goal, -->

<h1>1. Programmable Output System </h1> <p> The purpose of this system is to release output DNA in order. We propose two different approaches.<br><br>

<h2>1st Approach; Enzyme system</h2> <p>Enzyme System has three processes.</p> 1. Amplifying Process:The process in which DNA Polymerase amplifies KEY DNA. <br> 2. Releasing Process:The process in which KEY DNA releases the liposome. <br> 3. Renewing Process:The process in which restriction enzyme renews the 3’ end of the input-DNA sequence to get an output. <br> <img src="http://openwetware.org/images/9/9e/Fde1a0-01.png">

<h3>1.Amplifying process</h3>

<p> In this process, first, domains A, B, and C in the template combine with domains A, B, and C in the input respectively. Then, polymerase copies 5’ end of the template sequence and extends input sequence. After that, nickase cleaves the end of the copied domain. Then, polymerase works at the gap created by the nickase and push out the domain A. Repeating this process again and again, we amplify the domain A. (Fig.1) </p> <p> This DNA domain, we call it A1, becomes an input to following process 2 and 3. </p>

<img src="http://openwetware.org/images/c/c9/Figure-01.png"><br><br>

<!--

&#9312; Domains A0, B0, and C0 in the template combine with domains A0-, B0-, and C0- in the input respectively. Input-Template complex is created.<br> &#9313; Polymerase recognizes 3’ end of input. (Identified with dashed circle) <br> &#9314; Extending input sequence. The sequence A1 is newly formed. <br> &#9315; Polymerase runs away from Input-Template complex. <br> &#9316; Nickase combines with its recognition cite. <br> &#9317; Cleaving the end of the copied domain./ Making nick between A0 and A1. <br> &#9318; Polymerase works from the gap (Identified with dashed circle) created by the nickase. <br> &#9319; Running on DNA with making nucleotides and pushing out the A1. <br> &#9320; Repeating this process again and again amplifies the number of domain A1. <br><br>

-->

<h3>2.Releasing process</h3> <p> A1 combines with the 3’ end of the DNA which is combined with the DNA modified with liposome (output-A). Polymerase recognizes the 3’ end of the A1 and then extend the chain. </p> <p> At the same time, output-A is denatured and released. </p> <img src="http://openwetware.org/images/1/1f/IMG_3657-02.jpg" width="735px" height="600px"> <p>A1 combines with the end of the DNA which fixes the liposome-modifying DNA (output-A). Polymerases recognize the 3’end of the A1 and extend the chain. At the same time, output-A is denatured and released.</p>


<h3>3.Renewing process</h3> <img src="http://openwetware.org/images/7/7f/Figure3-01.png"> <p> When A1 combines with the gate-A, the DNA which has the recognition sequence corresponding to the restriction enzyme is released by polymerase. When this DNA combines with the structure made of input and template, a restriction enzyme activates, and cleaves the chain between A and B. By doing this, the domain near the 3’ end of the input becomes domain B. Then, the process goes back to the system-1. </p> <p> In this way, output-A, output-B, and output-C are released in order.</p>

<h2>2nd Approach; Enzyme-free System</h2> <img src="http://openwetware.org/images/9/97/Figure_EnzymeFree-01.png"> <img src="http://openwetware.org/images/f/f6/Figure_EnzymeFree-03.png"> <img src="http://openwetware.org/images/8/89/Figure_EnzymeFree-04.png"> <p>This approach is inspired by seesaw gate (Lulu Qian et.al, 2011). Our goal is to get different output signals in order of input signals. In this system , we should arrange a input , a trigger ,fuels, the double strand DNA bonding with liposome.</p> <img src="http://openwetware.org/images/7/79/Figure_EnzymeFree-02.png"> <p>（用意するもの） fuel,gate、Liposomeが結合した二本鎖DNA (Fa,b,c Ga,b,c La,b,c) input(I1),trigger(T1)それぞれ一種類 </p>

<p>Reactions as follows : </p> <h3>1.</h3> <p> Input-DNA sequence are dissolved in the solution including trigger , fuels , gates , and the double strand DNA bonding with liposome. Then, 　trigger combines with input and the single strand DNA (DNA1)which makes input are released. </p>

<h3>2.</h3> <p> DNA1 combines with the double strand DNA bonding with liposome . In this reaction , the single strand DNA bonding with liposome are released due to the difference in length of DNA sequence which makes the double strand DNA. This reaction causes earlier so that toehold is longer than that of gate. 

</p>

<h3>3.</h3> <p> Other DNA1 combine with gate(Ga). the single strand DNA (DNA2)which makes gate(Ga) are released so that the length of the double strand is difference. In addition , DNA1 combines with gate. </p>

<h3>4.</h3> <p> DNA1 is released again so that DNA3 react with fuel(Fa) , due to the difference in length of DNA. 

</p>

<h3>5.</h3> <p> When we perform process3 and 4 repeatedly , the number of DNA2 is increasing. DNA2 is the key of next process. </p> <img src="http://openwetware.org/images/4/42/Enzyme-free_System_picture2.jpg" width="441px" height="599px">

<h3>6.</h3> <p> DNA2 combines with input as trigger , so the reaction said above caused.In this reaction , DNA is released as DNA2 in the reaction　said above . </p> <img src="http://openwetware.org/images/2/20/Enzyme-free_System_picture3.jpg" width="426px" height="600px">

<h3>7.</h3> <p> DNA4 combines with input as trigger , so the reaction said above caused. </p> <img src="http://openwetware.org/images/d/d4/Enzyme-free_System_picture4.jpg" width=395px height="599px">

<p> We can get output signals in order by using this reaction. </p>

<h2>2. Taste releasing system </h2> <p> The purpose of this system is to encapsulate taste substances in liposome and to release liposome attached to substrate.

The DNA which has the complementary sequence (comp DNA) to the Output is sprouted on agarose gel. They are hybridized each other. When A1 reacts with comp DNA, the output is separated from comp DNA, then the liposome is released from the agarose gel, and taste substances are diffused. </p>

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