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| | <span style="line-height:46px;" class="line">Development of A Novel</span></span> | | <span style="line-height:46px;" class="line">Development of A Novel</span></span> |
| | <span style="line-height:46px;" class="line-wrap"> | | <span style="line-height:46px;" class="line-wrap"> |
| − | <span style="line-height:46px;" class="line">Blood-MicroRNA Rapid Detection System with CRISPR</span></span> | + | <span style="line-height:46px;" class="line">Blood-MicroRNA Handy Detection System with CRISPR</span></span> |
| | <!--</a>--></h2> | | <!--</a>--></h2> |
| | </div> | | </div> |
| Line 354: |
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| | <a style="color:rgb(10,31,84);" href="/Team:NUDT_CHINA">HomePage</a> • | | <a style="color:rgb(10,31,84);" href="/Team:NUDT_CHINA">HomePage</a> • |
| | <!-- 修改这里！！ -->AWARDS • | | <!-- 修改这里！！ -->AWARDS • |
| − | <a style="color:rgb(10,31,84);" href="/Team:NUDT_CHINA/Model"><!-- 修改这里！！ -->Model</a> | + | <a style="color:rgb(10,31,84);" href="/Team:NUDT_CHINA/Description"><!-- 修改这里！！ -->Model</a> |
| | </h5><hr style="width:40%;margin-left:60%;border-top:1px solid rgb(10,31,84);" /></div> | | </h5><hr style="width:40%;margin-left:60%;border-top:1px solid rgb(10,31,84);" /></div> |
| | | | |
| Line 377: |
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| | | | |
| | | | |
| − |
| + | <!--在这里插入内容--> |
| − | | + | |
| − | <!--标题--> | + | |
| − | <h2>
| + | |
| − | <span><span style="color:#7f1015">Abstract</span></span><hr />
| + | |
| − | </h2>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">This
| + | |
| − | | + | |
| − | model is created to evaluate the effectiveness of initial design, and offers
| + | |
| − | | + | |
| − | guidelines about how the system can (or must) be improved. (You can go<a
| + | |
| − | | + | |
| − | href="/Team:NUDT_CHINA/Design"> PROJECT.page </a>to see more.) The core idea is
| + | |
| − | | + | |
| − | to simulate the process of producing the
| + | |
| − | | + | |
| − | signal which can be detected, and draw a conclusion by obtaining the
| + | |
| − | | + | |
| − | relationship between the signal intensity and the concentration of miRNA.
| + | |
| − | | + | |
| − | </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h2>
| + | |
| − | <span><span style="color:#7f1015">Introduction </span></span><hr />
| + | |
| − | </h2>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | create mathematical models of two aspects of our project, a RCA model and a
| + | |
| − | | + | |
| − | signal detection model.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | RCA
| + | |
| − | | + | |
| − | model is based on the Michaelis-Menten equation. We can obtain the relationship
| + | |
| − | | + | |
| − | between the concentration of miRNA and the number of stem-loop structures
| + | |
| − | | + | |
| − | through theoretical calculation, and we use our experimental results to
| + | |
| − | | + | |
| − | calculate the parameters we introduced previously.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">In the
| + | |
| − | | + | |
| − | signal detection model, we combine a probability model with the kinetic
| + | |
| − | | + | |
| − | equation
| + | |
| − | | + | |
| − | of enzymatic reaction, so we can obtain the relationship between the number of
| + | |
| − | | + | |
| − | stem-loop structures and the signal intensity under the premise of adding a
| + | |
| − | | + | |
| − | certain amount of the fusion proteins of dCas9 and split-HRP fragments. </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">By
| + | |
| − | | + | |
| − | integrating the two models, we can theoretically predict the impacts of the
| + | |
| − | | + | |
| − | molecule number of proteins on the signal to noise ratio and explained the
| + | |
| − | | + | |
| − | trend
| + | |
| − | | + | |
| − | of the signal intensity with the change of the concentration of miRNA in our
| + | |
| − | | + | |
| − | wet-lab work.
| + | |
| − | </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h2>
| + | |
| − | <span><span style="color:#7f1015"> Assumption and Justification
| + | |
| − | | + | |
| − | </span></span><hr />
| + | |
| − | </h2>
| + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015"> About model </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | assume that miRNA is not degraded throughout the reaction process.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | assume that the two fusion proteins of dCas9 and split-HRP fragments have the
| + | |
| − | | + | |
| − | same ability to combine with the stem-loop structure, and only when two
| + | |
| − | | + | |
| − | different proteins next to each other, can they have the ability to catalyze
| + | |
| − | | + | |
| − | substrate and produce signal.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | assume that the number of stem-loop structures in each RCA product is equal
| + | |
| − | | + | |
| − | under a certain reaction time. </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | assume that the enzymatic activity remains unchanged with time under the
| + | |
| − | | + | |
| − | premise
| + | |
| − | | + | |
| − | of excessive amount of enzymes or a short-time reaction.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015"> About the data </span></span><hr
| + | |
| − | | + | |
| − | />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | assume that the data we obtain from wet-lab experiment are reliable.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We
| + | |
| − | | + | |
| − | assume that all the results are trustworthy in the process of statistical
| + | |
| − | | + | |
| − | processing and data calculation.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h2>
| + | |
| − | <span><span style="color:#7f1015">Model </span></span><hr />
| + | |
| − | </h2>
| + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015">Notations </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--表格-->
| + | |
| − | <table class="MsoTableGrid" border="1" cellspacing="0" cellpadding="0"
| + | |
| − | | + | |
| − | style="border:none;">
| + | |
| − | <tbody>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>Symbol </span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>Definition </span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>x</span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a name="OLE_LINK7"></a><a
| + | |
| − | | + | |
| − | name="OLE_LINK6"></a><span>The concentration of</span><span> miRNA (pM)</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>C<sub>1</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The concentration of initiated
| + | |
| − | | + | |
| − | probe (Abbreviated
| + | |
| − | to iprobe)</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>k<sub>1</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a name="OLE_LINK8"></a><span>A
| + | |
| − | | + | |
| − | constant
| + | |
| − | | + | |
| − | representing
| + | |
| − | the scale factor</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>K<sub>m </sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>One of the characteristic
| + | |
| − | | + | |
| − | constants of
| + | |
| − | phi29 DNA polymerase</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>V<sub>max</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>One of the characteristic
| + | |
| − | | + | |
| − | constants of phi29
| + | |
| − | DNA polymerase</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>k<sub>2</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>A constant representing the scale
| + | |
| − | | + | |
| − | factor</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>V</span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The initial speed of RCA </span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a
| + | |
| − | | + | |
| − | name="OLE_LINK9"></a><i><span>n<sub>1</sub></span></i><i><sub><span></span></su
| + | |
| − | | + | |
| − | b
| + | |
| − | | + | |
| − | ></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The moles of RCA product</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>n<sub>2</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a name="OLE_LINK10"></a><span>The
| + | |
| − | | + | |
| − | number of
| + | |
| − | stem-loop structures in each RCA product</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>n</span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The total amount of <a
| + | |
| − | | + | |
| − | name="OLE_LINK23"></a><a name="OLE_LINK22"></a>stem-loop structures</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>N</span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a name="OLE_LINK41"></a><a
| + | |
| − | | + | |
| − | name="OLE_LINK16"></a><span>The molecule number of the
| + | |
| − | fusion protein of dCas9 and split-HRP fragments</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>k<sub>3</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>A constant representing the scale
| + | |
| − | | + | |
| − | factor</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>y<sub>1</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The fluorescence intensity of
| + | |
| − | DNA-dye-complex (RFU)</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>N<sub>1</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The molecule number of the fusion
| + | |
| − | | + | |
| − | protein
| + | |
| − | of dCas9 and split-HRP fragments in the solution</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>N<sub>2</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a name="OLE_LINK45"></a><span>The
| + | |
| − | | + | |
| − | molecule number
| + | |
| − | of the fusion protein of dCas9 and split-HRP fragments binding with stem-loop
| + | |
| − | structure</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>k<sub>4</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>A constant representing the scale
| + | |
| − | | + | |
| − | factor</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span>k<sub>5</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>A constant representing the scale
| + | |
| − | | + | |
| − | factor</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span style="font-family:宋
| + | |
| − | | + | |
| − | 体;">ρ</span><span></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>Signal to noise ratio(Abbreviated
| + | |
| − | | + | |
| − | to SNR)</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span style="font-family:宋
| + | |
| − | | + | |
| − | 体;">I</span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The molecule number of formed
| + | |
| − | | + | |
| − | intact HRP
| + | |
| − | proteins </span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span style="font-family:宋
| + | |
| − | | + | |
| − | 体;">I<sub>1</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The molecule number of formed
| + | |
| − | | + | |
| − | intact HRP
| + | |
| − | proteins in the solution</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span style="font-family:宋
| + | |
| − | | + | |
| − | 体;">I<sub>2</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <a name="OLE_LINK43"></a><span>The
| + | |
| − | | + | |
| − | molecule number
| + | |
| − | of formed intact HRP proteins through binding with stem-loop structure</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span style="font-family:宋
| + | |
| − | | + | |
| − | 体;">t</span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>Reaction time</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td width="66" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <i><span style="font-family:宋
| + | |
| − | | + | |
| − | 体;">y<sub>2</sub></span></i>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | <td width="487" valign="top" style="border:solid
| + | |
| − | | + | |
| − | windowtext 1.0pt;">
| + | |
| − | <p class="MsoNormal">
| + | |
| − | <span>The signal intensity
| + | |
| − | | + | |
| − | (OD<sub>450</sub>)</span>
| + | |
| − | </p>
| + | |
| − | </td>
| + | |
| − | </tr>
| + | |
| − | </tbody>
| + | |
| − | </table>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015"> The RCA model </span></span><hr
| + | |
| − | | + | |
| − | />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Firstly, we
| + | |
| − | | + | |
| − | assume that there is a linear relationship between the
| + | |
| − | | + | |
| − | concentration of the initiated probes and miRNA as the combination reaction
| + | |
| − | | + | |
| − | between miRNA and probe occurs spontaneously<sup>1 </sup> . This can be written
| + | |
| − | | + | |
| − | as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model1.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Next,
| + | |
| − | | + | |
| − | RCA is an enzymatic reaction, the reaction equation is written as: iprobe+dNTP
| + | |
| − | | + | |
| − | →
| + | |
| − | | + | |
| − | iprobe-dN+PPi, under the premise of excessive amount of enzymes and dNTPs, the
| + | |
| − | | + | |
| − | relationship between the concentration of iprobe and the initial speed of RCA
| + | |
| − | | + | |
| − | can be described by the Michaelis-Menten equation. This can be written as：
| + | |
| − | | + | |
| − | </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model2.jpg|105px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">It is
| + | |
| − | | + | |
| − | obvious that there is a linear relationship between the molecule numbers of
| + | |
| − | | + | |
| − | iprobe-dN (n<sub>1 </sub>
| + | |
| − | , reflects the molecule numbers of RCA product) and the initial speed of RCA
| + | |
| − | | + | |
| − | reaction. Notably, under the premise of excessive amount of enzymes, the
| + | |
| − | | + | |
| − | extending of each RCA production is related to the reaction time, not the
| + | |
| − | | + | |
| − | concentration of the iprobe. It is a linear relationship when we assume that
| + | |
| − | | + | |
| − | the
| + | |
| − | | + | |
| − | enzymatic activity remains unchanged with time, thus, the number of stem-loop
| + | |
| − | | + | |
| − | structures (n<sub>2 </sub>
| + | |
| − | ) in each RCA product is stable under the premise of a certain reaction time.
| + | |
| − | | + | |
| − | </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig1.jpg|500px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure 1. Schematic diagram</b></span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">This
| + | |
| − | | + | |
| − | can be written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model3.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model4.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">And
| + | |
| − | | + | |
| − | then, SYBR Green binds to DNA. The
| + | |
| − | resulting DNA-dye-complex absorbs blue light (<i>λ<sub>max</sub></i> = 497nm)
| + | |
| − | | + | |
| − | and emits green light (<i>λ<sub>max</sub></i> = 520nm). Thus, there is
| + | |
| − | a linear relationship between the total amount of stem-loop structures and the
| + | |
| − | | + | |
| − | fluorescence
| + | |
| − | intensity of DNA-dye-complex. This can be written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model5.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">In
| + | |
| − | | + | |
| − | summary, the relationship between the concentration of miRNA and the
| + | |
| − | | + | |
| − | fluorescence intensity of DNA-dye-complex can be written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model6.jpg|210px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015"> The signal detection model
| + | |
| − | | + | |
| − | </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Firstly, it is
| + | |
| − | | + | |
| − | obvious that there is a linear relationship between
| + | |
| − | | + | |
| − | the molecule number of formed intact HRP proteins in the solution (consider as
| + | |
| − | | + | |
| − | NOISE) and the molecule number of the fusion proteins in the solution. This can
| + | |
| − | | + | |
| − | be written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model7.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Next,
| + | |
| − | | + | |
| − | we build a probability model to find the expression of the molecule number of
| + | |
| − | | + | |
| − | formed intact HRP proteins (fused with dCas9) through binding with stem-loop
| + | |
| − | | + | |
| − | structure (consider as SIGNAL). It is related to the total amount of stem-loop
| + | |
| − | | + | |
| − | structures and the molecule number of DNA-bound fusion proteins. We obtain the
| + | |
| − | | + | |
| − | result by Monte Carlo method. </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig2.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure2. Frequency Distribution</b> Plot of frequency
| + | |
| − | | + | |
| − | distribution of the molecule number of formed intact HRP proteins through
| + | |
| − | | + | |
| − | binding with stem-loop structure. </span>
| + | |
| − | | + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig3.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure 3. The result of the probability model</b> Plot of the
| + | |
| − | | + | |
| − | expect value of the molecule number of formed intact proteins against the total
| + | |
| − | | + | |
| − | amount of stem-loop structures, carried out at 0.1, 0.3, 0.5, 0.7, 0.9 of
| + | |
| − | | + | |
| − | N<sub>2</sub> to n ratio.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | result can be written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model8.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">As for
| + | |
| − | | + | |
| − | <i>N<sub>2</sub></i>,
| + | |
| − | which refers to the molecule number of the fusion proteins binding with
| + | |
| − | stem-loop structure. The equation is formulated based on the limiting case. The
| + | |
| − | equation can be written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model9.jpg|105px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">And
| + | |
| − | | + | |
| − | then, we can obtain the expressions for <i>ρ</i></span> <span>and<i> I</i>
| + | |
| − | | + | |
| − | separately.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model10.jpg|50px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model11.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Notably, under
| + | |
| − | | + | |
| − | the premise of excessive substrates,
| + | |
| − | it is a linear relationship between the reaction rate and the concentration of
| + | |
| − | enzymes when we assume that the enzymatic activity remain unchanged with time,
| + | |
| − | thus, we can obtain the expression of the signal intensity at <i>t<sub>th</sub>
| + | |
| − | | + | |
| − | </i>time-step. This is
| + | |
| − | written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model12.jpg|70px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">In
| + | |
| − | | + | |
| − | summary, the relationship between the number of stem-loop structures and SNR
| + | |
| − | | + | |
| − | under the premise of adding a certain amount of the fusion proteins can be
| + | |
| − | | + | |
| − | written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model13.jpg|140px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | relationship between the number of Stem-loop structures and the signal
| + | |
| − | | + | |
| − | intensity
| + | |
| − | | + | |
| − | under the premise of adding a certain amount of the fusion proteins can be
| + | |
| − | | + | |
| − | written as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model14.jpg|280px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015"> The calculation of the constants
| + | |
| − | | + | |
| − |
| + | |
| − | | + | |
| − | </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">To
| + | |
| − | | + | |
| − | simplify the equation (1), the constants
| + | |
| − | can be integrated. As can be seen from the above table, <i>k<sub>1</sub></i>,
| + | |
| − | | + | |
| − | <i>k<sub>2</sub></i>, <i>k<sub>3</sub></i><sub> </sub>are
| + | |
| − | constants representing the scale factor, <i>K<sub>m</sub></i> and
| + | |
| − | | + | |
| − | <i>V<sub>max</sub></i> are
| + | |
| − | characteristic constants of phi 29 DNA polymerase, and <i>n<sub>2</sub></i>,
| + | |
| − | | + | |
| − | which refers to the number of stem-loop structures
| + | |
| − | in each RCA product, is stable under the premise of a certain reaction time.
| + | |
| − | Thus, define two constants, mark as <i>a</i> and <i>b</i>, and then the
| + | |
| − | | + | |
| − | equation
| + | |
| − | | + | |
| − | can be
| + | |
| − | simplified as:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--插入图片-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--model15.jpg|105px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We use
| + | |
| − | | + | |
| − | this equation to fit the data points obtained through experiments. (Figure 2.
| + | |
| − | | + | |
| − | (D) in the<a href="/Team:NUDT_CHINA/Results"> RESULTS.page. </a>) The fitting
| + | |
| − | | + | |
| − | curve is shown below.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig4.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure 4. Fitting curve</b> Fitting curve of fluorescence
| + | |
| − | | + | |
| − | intensity of DNA-dye-complex (RFU) against the concentrations of miRNA
| + | |
| − | | + | |
| − | (pM).</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">With
| + | |
| − | | + | |
| − | regard to the equation (2), we set the parameters based on the experiments as
| + | |
| − | | + | |
| − | follows:
| + | |
| − | <i>k<sub>3</sub> </i>=8.35*10<sup>-16 </sup>
| + | |
| − | | + | |
| − | (<i>n</i>=<i>y<sub>1</sub></i>/<i>k<sub>3</sub></i>); <i>k<sub>4</sub></i>
| + | |
| − | | + | |
| − | =0.038;<i> k<sub>5</sub>t</i>=5.9*10<sup>-12</sup>.
| + | |
| − | </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015"> Results and Analysis
| + | |
| − | | + | |
| − | </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">By
| + | |
| − | | + | |
| − | integrating the two models, we can obtain the relationships between SNR, the
| + | |
| − | | + | |
| − | signal intensity respectively and the concentration of miRNA under different
| + | |
| − | | + | |
| − | addition amount of fusion proteins.
| + | |
| − | </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | results are shown below.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig5.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure
| + | |
| − | 5. The signal intensity (OD<sub>450</sub>)</b> Three-dimensional map of signal
| + | |
| − | | + | |
| − | intensity (OD<sub>450</sub>)
| + | |
| − | against miRNA concentration(pM) and additional amount of fusion
| + | |
| − | | + | |
| − | proteins.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">There
| + | |
| − | | + | |
| − | exists a monotonous relation between the concentration of miRNA and the signal
| + | |
| − | | + | |
| − | intensity when the value of the molecule number of the fusion proteins is
| + | |
| − | | + | |
| − | relatively large. While the relationship does not hold when the value of the
| + | |
| − | | + | |
| − | molecule number of the fusion proteins is relatively small. And the signal
| + | |
| − | | + | |
| − | intensity increases as the value of the molecule number of the fusion proteins
| + | |
| − | | + | |
| − | increases.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig6.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure 6. The result of SNR</b> Three-dimensional map of signal
| + | |
| − | | + | |
| − | to noise ratio against miRNA concentration (pM) and additional amount of fusion
| + | |
| − | | + | |
| − | proteins.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | signal-to-noise ratio decreases as the value of the molecule number of fusion
| + | |
| − | | + | |
| − | proteins increases. </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015">Test of model </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Our
| + | |
| − | | + | |
| − | model predicts the relationship between the signal intensity and the
| + | |
| − | | + | |
| − | concentration of miRNA under the premise of adding a certain amount of the
| + | |
| − | | + | |
| − | fusion proteins of dCas9 and split-HRP fragments. So we can test if our model
| + | |
| − | | + | |
| − | is reasonable by comparing the curves predicted by the model with those
| + | |
| − | | + | |
| − | obtained from the experiment. The results are shown below.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig7-1.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig7-2.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure 7. Test of model</b> (A) Plot of the signal
| + | |
| − | intensity (OD<sub>450</sub>) against miRNA concentration (pM) predicted by the
| + | |
| − | model with the value of the molecule number of the fusion proteins set to be
| + | |
| − | | + | |
| − | 4.66e+13.
| + | |
| − | (B)<span> Plot of the signal
| + | |
| − | intensity (OD<sub>450</sub>) against miRNA concentration (pM) obtained from the
| + | |
| − | experiment with the value of the molecule number of the fusion proteins set to
| + | |
| − | be 4.66e+13. (C) Plot of the signal intensity
| + | |
| − | (OD<sub>450</sub>) against miRNA concentration (pM) predicted by the model with
| + | |
| − | the value of the molecule number of the fusion proteins set to be 4.66e+14. (D)
| + | |
| − | Plot of the signal intensity (OD450) against miRNA concentration (pM) obtained
| + | |
| − | from the experiment with the value of the molecule number of the fusion
| + | |
| − | proteins set to be 4.66e+14.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">We can
| + | |
| − | | + | |
| − | learn from the figure above that the curves predicted by the model are in good
| + | |
| − | | + | |
| − | agreement with the experimental results.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h3>
| + | |
| − | <span><span style="color:#7f1015">Model extension </span></span><hr />
| + | |
| − | </h3>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Furthermore,
| + | |
| − | | + | |
| − | we consider that it is conducive to signal detection when the value of signal
| + | |
| − | | + | |
| − | intensity and SNR are both greater than two. We obtain the ideal region through
| + | |
| − | | + | |
| − | calculation, which is shown as below. </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--图和图注-->
| + | |
| − | </br>
| + | |
| − | </html>
| + | |
| − | [[File:T--NUDT_CHINA--modelfig7.jpg|700px|center]]
| + | |
| − | <html>
| + | |
| − | </br>
| + | |
| − | <p>
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-
| + | |
| − | | + | |
| − | size:18px;"><b>Figure 8. Ideal region</b> Region of qualified logarithm of
| + | |
| − | | + | |
| − | concentrations of miRNA (pM) and logarithm of the additional amount of fusion
| + | |
| − | | + | |
| − | proteins.</span>
| + | |
| − | </p>
| + | |
| − | | + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">When
| + | |
| − | | + | |
| − | the
| + | |
| − | value of <i>N</i> is set to e+13.3, the range of the value of<i> x</i>
| + | |
| − | | + | |
| − | contained in the ideal region is<span> maximum, which means when the value of
| + | |
| − | | + | |
| − | the
| + | |
| − | molecule number of the fusion proteins is e+13.3, the concentration range of
| + | |
| − | miRNA that can be detected is maximum. </span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h2>
| + | |
| − | <span><span style="color:#7f1015">Conclusion </span></span><hr />
| + | |
| − | </h2>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">Based
| + | |
| − | | + | |
| − | on our simulation, we came to the conclusion that:</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | curves predicted by the model are in good agreement with the experimental
| + | |
| − | | + | |
| − | results, thus indicating that our model is reasonable.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | value of the molecule number of the fusion protein of dCas9 and split-HRP
| + | |
| − | | + | |
| − | fragments not only affects the signal-to-noise ratio, but also the signal
| + | |
| − | | + | |
| − | intensity. So we need to weigh its impact on both to select the optimal
| + | |
| − | | + | |
| − | solution.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p style="text-indent:22pt;">
| + | |
| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">The
| + | |
| − | | + | |
| − | concentration range of miRNA that can be detected is maximum when we set the
| + | |
| − | | + | |
| − | value of the molecule number of the fusion proteins to be e+13.3, building on
| + | |
| − | | + | |
| − | which, our wet-lab protocol could be optimized in our future work.</span>
| + | |
| − | </p>
| + | |
| − | </br>
| + | |
| − | | + | |
| − | | + | |
| − | <!--标题-->
| + | |
| − | <h2>
| + | |
| − | <span><span style="color:#7f1015">Reference </span></span><hr />
| + | |
| − | </h2>
| + | |
| − | | + | |
| − | <!--正文-->
| + | |
| − | <p >
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| − | <span style="line-height:2;font-family:Perpetua;font-size:18px;">1 Deng, R. <i>et al</i>. Toehold-initiated rolling circle amplification for visualizing individual microRNAs in situ in single cells. <i>Angew Chem Int Ed Engl </i>53, 2389-2393,doi:10.1002/anie.201309388 (2014).</span>
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