Difference between revisions of "Team:BGIC-Union/Model"

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 +
 
<html>
 
<html>
 +
  <head>
 +
  <html>
 
   <head>
 
   <head>
 
     <meta charset="utf-8">
 
     <meta charset="utf-8">
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−
     <title>Team：BGIC-Union/MODEL</title>
+
     <title>Team：BGIC-Union/Practices-Design</title>
  
 
   <head>
 
   <head>
 
    
 
    
−
        <link href='http://fonts.googleapis.com/css?family=Lato:400,300' rel='stylesheet' type='text/css'>
 
−
        <link href='http://fonts.googleapis.com/css?family=Raleway:400,300,500' rel='stylesheet' type='text/css'>
 
  
−
<!-- progress -->
+
   <!-- Bootstrap core CSS -->
−
<link href="https://2017.igem.org/Team:BGIC-Union/mainstyle!/css?action=raw&amp;ctype=text/css" rel="stylesheet">
+
−
    
+
−
<!-- Bootstrap core CSS -->
+
 
     <link href="https://2017.igem.org/Template:BGIC-Union/bootstrap-css?action=raw&amp;ctype=text/css" rel="stylesheet">
 
     <link href="https://2017.igem.org/Template:BGIC-Union/bootstrap-css?action=raw&amp;ctype=text/css" rel="stylesheet">
 +
<!-- animate -->
 +
<link href="https://2017.igem.org/Template:BGIC-Union/animate-css?action=raw&amp;ctype=text/css" rel="stylesheet">
 +
  <!-- carousel -->
 +
  <link href="https://2017.igem.org/Template:BGIC-Union/owlcarousel-css?action=raw&amp;ctype=text/css" rel="stylesheet">
 +
  <link href=" https://2017.igem.org/Template:BGIC-Union/fontawesomemin2-css?action=raw&amp;ctype=text/css" rel="stylesheet">
 +
  <!-- light -->
 +
<style type="text/css">
 +
/* fixed line holder */
 +
.timelineLight .timeline_line {c
 +
margin-top:50px;
 +
margin-bottom:10px;
 +
width:900px;
 +
}
 +
.timelineLight
 +
{color: #fff;
 +
background: #ffe4e1;}
 +
/* full (including months that are not shown) line holder */
 +
.timelineLight .t_line_holder {
 +
height:80px;
 +
background:url('https://static.igem.org/mediawiki/2017/3/35/Line.jpg') repeat-x 0px 39px;
 +
}
  
 +
/* 2 months are stored in one view */
 +
.timelineLight .t_line_view {
 +
height:20px;
 +
width:900px;
 +
}
 +
 +
/* holder for year number */
 +
.timelineLight h3.t_line_year {
 +
margin:0;
 +
color:#545454;
 +
}
 +
 +
/* holder for 1 month (constist of nodes and month caption) - we use borders to separate months thats why it has width 2px less then 50% */
 +
.timelineLight .t_line_m {
 +
margin-top:35px;
 +
height:10px;
 +
border-left:1px solid #545454;
 +
border-right:1px solid #545454;
 +
width:448px;
 +
}
 +
 +
/* month on the right side - has left set at 459 so border would overlap border from first element (to evade duplicated borders) */
 +
.timelineLight .t_line_m.right {
 +
left:449px;
 +
width:449px;
 +
}
 +
 +
/* month caption */
 +
.timelineLight h4.t_line_month {
 +
margin:-30px 0 0;
 +
color:#545454;
 +
}
 +
 +
/* used in responsive layout when only one month is shown (it is span containing year) */
 +
.t_line_month_year {
 +
display:none;
 +
}
 +
 +
/* node on the timeline */
 +
.timelineLight a.t_line_node {
 +
text-decoration:none;
 +
padding:38px 0 4px;
 +
height:10px;
 +
font-size:12px;
 +
top:-25px;
 +
background:url('https://static.igem.org/mediawiki/2017/2/2a/Dot.png') no-repeat center 24px;
 +
color:#141817;
 +
}
 +
.timelineLight a.t_line_node:hover {
 +
background:url('https://static.igem.org/mediawiki/2017/6/65/Dot-rollover.png') no-repeat center 24px;
 +
}
 +
.timelineLight a.t_line_node.active {
 +
background:url('https://static.igem.org/mediawiki/2017/d/d7/Dot-selected.png') no-repeat center 24px;
 +
}
 +
 +
/* node description */
 +
.timelineLight .t_node_desc {
 +
background: rgba(26,134,172,0.9);
 +
top:0;
 +
color:#fff;
 +
padding:1px 5px;
 +
}
 +
 +
/* descriptions on right side go from right to left */
 +
.timelineLight .t_node_desc.pos_right {
 +
right:0;
 +
}
 +
 +
/* line arrow left */
 +
.timelineLight #t_line_left {
 +
cursor:pointer;
 +
left:-30px;
 +
top:30px;
 +
width:14px;
 +
height:19px;
 +
background:url('https://static.igem.org/mediawiki/2017/9/95/Bgic-arrow.png') no-repeat left top;
 +
}
 +
 +
.timelineLight #t_line_left:hover {
 +
background:url('https://static.igem.org/mediawiki/2017/9/95/Bgic-arrow.png') no-repeat left bottom;
 +
}
 +
 +
/* line arrow right */
 +
.timelineLight #t_line_right {
 +
cursor:pointer;
 +
right:-30px;
 +
top:30px;
 +
width:14px;
 +
height:19px;
 +
background:url('https://static.igem.org/mediawiki/2017/9/95/Bgic-arrow.png') no-repeat right top;
 +
}
 +
 +
.timelineLight #t_line_right:hover {
 +
background:url('https://static.igem.org/mediawiki/2017/9/95/Bgic-arrow.png') no-repeat right bottom;
 +
}
 +
.timelineLight {
 +
display:none;
 +
}
 +
 +
/* items container */
 +
.timelineLight .timeline_items {
 +
padding:10px 0;
 +
}
 +
 +
/* single item (margines set from javascript) */
 +
.timelineLight .item {
 +
height:380px;
 +
text-align:center;
 +
 +
color: #fff;
 +
background: #f7dae7;
 +
 +
-moz-box-shadow: -3px 1px 6px rgba(0,0,0,0.4);
 +
-webkit-box-shadow: -3px 1px 6px rgba(0,0,0,0.4);
 +
box-shadow: -3px 1px 6px rgba(0,0,0,0.4);
 +
width:200px;
 +
}
 +
 +
/* ----- content - non-plugin elements ----- */
 +
.timelineLight .item img {
 +
margin-top:0;
 +
width:200px;
 +
}
 +
 +
.timelineLight .item span{
 +
display:block;
 +
margin:0px 20px 10px;
 +
height:150px;
 +
}
 +
.timelineLight .item .read_more {
 +
padding:2px 8px 2px 10px;
 +
font-family:Comic Sans MS;
 +
font-size:20px;
 +
float:right;
 +
color:#ffffff;
 +
background: rgba(0,0,0,0.35);
 +
cursor:pointer;
 +
}
 +
.timelineLight .item .read_more:hover {
 +
background:rgb(26,134,172);
 +
}
 +
/* ----------------------------------------- */
 +
 +
/* item details (margines set from javascript) */
 +
.timelineLight .item_open {
 +
height:380px;
 +
background:url('../images/timeline/light/background.jpg') repeat;
 +
position:relative;
 +
color:#545454;
 +
z-index:2;
 +
-moz-box-shadow: 0px 0px 6px rgba(0,0,0,0.4);
 +
-webkit-box-shadow: 0px 0px 6px rgba(0,0,0,0.4);
 +
box-shadow: 0px 0px 6px rgba(0,0,0,0.4);
 +
width:490px;
 +
 +
}
 +
 +
/* item details content wrapper (used for animation - shuld have same width as .item_open) */
 +
.timelineLight .item_open_cwrapper {
 +
width:490px;
 +
}
 +
 +
/* ----- content - non-plugin elements ----- */
 +
.timelineLight .timeline_open_content {
 +
padding:20px;
 +
}
 +
 +
.timelineLight .item_open h2 {
 +
margin-top:10px;
 +
padding-top:0;
 +
font-size:28px;
 +
}
 +
.timelineLight .item_open .t_close {
 +
position:absolute;
 +
top:10px;
 +
right:10px;
 +
padding:2px 8px 2px 10px;
 +
font-family: Comic Sans MS;
 +
font-size:20px;
 +
color:#ffffff;
 +
background: rgba(0,0,0,0.25);
 +
cursor:pointer;
 +
}
 +
.timelineLight .item_open .t_close:hover {
 +
background:rgb(26,134,172);
 +
}
 +
/* ----------------------------------------- */
 +
 +
/* left/right controles */
 +
.timelineLight .t_controles {
 +
margin:10px auto;
 +
text-align:center;
 +
}
 +
.timelineLight .t_left,
 +
.timelineLight .t_right {
 +
display:inline-block;
 +
height:50px;
 +
width:29px;
 +
margin:10px;
 +
cursor:pointer;
 +
}
 +
.timelineLight .t_left, 
 +
.timelineLight .t_left:hover:active {
 +
background: url('https://static.igem.org/mediawiki/2017/b/ba/Big-arrow.png') no-repeat left top;
 +
}
 +
.timelineLight .t_left:hover {
 +
background: url('https://static.igem.org/mediawiki/2017/b/ba/Big-arrow.png') no-repeat left bottom;
 +
}
 +
 +
.timelineLight .t_right,
 +
.timelineLight .t_right:hover:active{
 +
background: url('https://static.igem.org/mediawiki/2017/b/ba/Big-arrow.png') no-repeat right top;
 +
}
 +
 +
.timelineLight .t_right:hover {
 +
background: url('https://static.igem.org/mediawiki/2017/b/ba/Big-arrow.png') no-repeat right bottom;
 +
}
 +
 +
/* -----------------------------------------------------------------------*/
 +
/* ------------------------------ RESPONSIVE -----------------------------*/
 +
/* -----------------------------------------------------------------------*/
 +
 +
 +
 +
/* --- 768px --- */
 +
@media screen and (max-width:980px) {
 +
 +
.timelineLight .timeline_line {
 +
width:680px;
 +
}
 +
 +
.timelineLight .t_line_view {
 +
width:680px;
 +
}
 +
 +
.timelineLight .t_line_m {
 +
width: 338px;
 +
}
 +
.timelineLight .t_line_m.right {
 +
left: 339px;
 +
width: 339px;
 +
}
 +
}
 +
 +
 +
/* --- 610px --- */
 +
@media screen and (max-width:767px) {
 +
 +
.timelineLight .timeline_line {
 +
width:530px;
 +
}
 +
 +
.timelineLight .t_line_view {
 +
width:1060px;
 +
}
 +
 +
.timelineLight .t_line_m {
 +
width: 528px;
 +
}
 +
.timelineLight .t_line_m.right {
 +
left: 530px;
 +
width: 528px;
 +
}
 +
.timelineLight .t_line_year {
 +
opacity:0;
 +
filter:alpha(opacity=0)
 +
}
 +
.timelineLight .t_line_month_year {
 +
display:inline;
 +
}
 +
 +
.timelineLight .t_line_node span {
 +
 +
}
 +
.timelineLight .t_node_desc {
 +
font-size:8px;
 +
}
 +
.timelineLight .t_node_desc.pos_right {
 +
right:auto;
 +
left:0;
 +
}
 +
}
 +
 +
 +
/* --- 300px --- */
 +
@media screen and (max-width:
 +
 +
) {
 +
.timelineLight .timeline_line {
 +
width:240px;
 +
}
 +
 +
.timelineLight .t_line_view {
 +
width:480px;
 +
}
 +
 +
.timelineLight .t_line_m {
 +
width: 238px;
 +
}
 +
.timelineLight .t_line_m.right {
 +
left: 240px;
 +
width: 238px;
 +
}
 +
 +
.timelineLight .item_open {
 +
width:280px;
 +
height:390px;
 +
}
 +
.timelineLight .item_open_cwrapper {
 +
width:280px;
 +
}
 +
}
 +
 +
 +
 +
 +
</style>
 +
 +
  <!-- main -->
 +
  <style type="text/css">
 +
 
 +
 +
#top_title,#sideMenu { display: none !important; }
 +
        #content {
 +
            width: 100%;
 +
            margin: 0;
 +
            padding: 0;
 +
           
 +
        }
 +
 +
#bodyContent
 +
{
 +
margin-top：0px !important;
 +
}
 +
 +
 +
#content{
 +
  background-color: #FFe4e1; !important;
 +
}
 +
 +
 +
.mainstyle
 +
{
 +
font-family: Comic Sans MS;
 +
}
 +
html, body {
 +
    margin: 0 ;
 +
    padding: 0 ;
 +
}
 +
body {
 +
  background-color: #ffa07a;
 +
 
 +
    font-weight: 400;
 +
    font-size: 14px;
 +
    color: #555;
 +
    ；
 +
    -webkit-font-smoothing: antialiased;
 +
    -webkit-overflow-scrolling: touch;
 +
}
 +
 +
/* Titles */
 +
h1, h2, h3, h4, h5, h6 {
 +
font-family: Comic Sans MS;
 +
    font-weight: 700;
 +
    color: #333;
 +
}
 +
 +
h1 {
 +
font-size: 35px;
 +
margin-top: 30px;
 +
margin-bottom: 30px;
 +
}
 +
 +
 +
 +
 +
p {
 +
    line-height: 28px;
 +
    margin-bottom: 25px;
 +
    font-size: 16px;
 +
font-family: Comic Sans MS;
 +
}
 +
 +
.centered {
 +
    text-align: center;
 +
}
 +
 +
/* Links */
 +
a {
 +
    color: #20B2AA;
 +
    word-wrap: break-word;
 +
 +
    -webkit-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    -moz-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    -ms-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    -o-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    transition: color 0.1s ease-in, background 0.1s ease-in;
 +
}
 +
 +
a:hover,
 +
a:focus {
 +
    color: #7b7b7b;
 +
    text-decoration: none;
 +
    outline: 0;
 +
}
 +
 +
a:before,
 +
a:after {
 +
    -webkit-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    -moz-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    -ms-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    -o-transition: color 0.1s ease-in, background 0.1s ease-in;
 +
    transition: color 0.1s ease-in, background 0.1s ease-in;
 +
}
 +
 +
hr {
 +
    display: block;
 +
    height: 1px;
 +
    border: 0;
 +
    border-top: 1px solid #ccc;
 +
    margin: 1em 0;
 +
    padding: 0;
 +
}
 +
 +
.copyrights{
 +
text-indent:-9999px;
 +
height:0;
 +
line-height:0;
 +
font-size:0;
 +
overflow:hidden;
 +
}
 +
 +
.navbar {
 +
text-transform: uppercase;
 +
margin-bottom: 0px;
 +
}
 +
 +
.navbar-inverse {
 +
padding-bottom: 30px;
 +
padding-top: 0;
 +
}
 +
 +
.navbar-inverse {
 +
background-color: #FFA07A;
 +
border-color: #FA8072;
 +
}
 +
 +
.navbar-inverse .navbar-nav > li > a {
 +
color: white;
 +
 +
}
 +
 +
 +
.navbar-inverse .navbar-nav > li > a:hover {
 +
color: #222222;
 +
}
 +
 +
.navbar-brand {
 +
font-weight: 700;
 +
font-size: 40px;
 +
letter-spacing: 2px;
 +
backgroud: white;
 +
}
 +
 +
.navbar-inverse .navbar-brand {
 +
color: white;
 +
background-color: #FFA07A;
 +
 +
}
 +
 +
.navbar-inverse .navbar-toggle {
 +
border-color: transparent;
 +
font-size: 12px;
 +
}
 +
 +
.progress-bar-theme {
 +
  background-color: #FFA07A;
 +
}
 +
 +
 +
/* Helpers */
 +
 +
.mt {
 +
margin-top: 40px;
 +
margin-bottom: 40px;
 +
}
 +
 +
.pt {
 +
padding-top: 50px;
 +
padding-bottom: 50px;
 +
}
 +
 +
 +
/* +++++ WRAP SECTIONS +++++ */
 +
 +
#ww {
 +
padding-top: 0px;
 +
padding-bottom: 70px;
 +
background-color: #f2f2f2;
 +
}
 +
 +
 +
#footer {
 +
background-color: #ADD8E6;
 +
padding-top: 50px;
 +
padding-bottom: 50px;
 +
}
 +
 +
#footer p {
 +
color: white;
 +
font-family: Comic Sans MS;
 +
}
 +
 +
#footer h4 {
 +
color: white;
 +
text-transform: uppercase;
 +
padding-bottom: 20px;
 +
}
 +
 +
 +
/* +++++ Color Wraps / Blog Page +++++ */
 +
 +
#grey {
 +
padding-top: 60px;
 +
padding-bottom: 60px;
 +
background-color: #f2f2f2;
 +
}
 +
 +
#white {
 +
padding-top: 60px;
 +
padding-bottom: 60px;
 +
background-color: #ffffff;
 +
}
 +
 +
 +
 +
/* Blog Date*/
 +
bd {
 +
font-size: 12px;
 +
text-transform: uppercase;
 +
color: #d2d2d2;
 +
font-weight: 700;
 +
}
 +
 +
/* Blog Author*/
 +
ba {
 +
font-size: 12px;
 +
text-transform: uppercase;
 +
}
 +
 +
/* Blog Quote */
 +
.bq {
 +
font-size: 22px;
 +
padding-top: 30px;
 +
}
 +
 +
/* Blog Tags */
 +
bt {
 +
font-size: 12px;
 +
}
 +
 +
/* ================== PORTFOLIO IMAGES HOVER EFFECT ================== */
 +
/* Effects also are controled by hover.zoom.js */
 +
.zoom {
 +
    display:block;
 +
    position:relative;
 +
    overflow:hidden;
 +
    background:transparent url(https://static.igem.org/mediawiki/2017/f/fe/Bgic_loader.gif) no-repeat center;
 +
}
 +
 +
 +
    .zoomOverlay {
 +
        position:absolute;
 +
        top:0; left:0;
 +
        bottom:0; right:0;
 +
        display:none;
 +
        background-image:url(../img/zoom.png);
 +
        background-repeat:no-repeat;
 +
        background-position:center;
 +
    }
 +
 +
.zoom2 {
 +
opacity: 1;
 +
}
 +
 +
.zoom2:hover {
 +
opacity: 0.5;
 +
}
 +
 +
/* glyphicon Configuration */
 +
 +
.glyphicon {
 +
font-size: 40px;
 +
color: #1abc9c;
 +
}
 +
  </style>
 +
 
 +
  <!-- script -->
 +
  <script src="https://code.jquery.com/jquery-1.10.2.min.js"></script>
 +
  <script type="text/javascript">
 +
  /*
 +
* jQuery Easing v1.3 - http://gsgd.co.uk/sandbox/jquery/easing/
 +
*
 +
* Uses the built in easing capabilities added In jQuery 1.1
 +
* to offer multiple easing options
 +
*
 +
* TERMS OF USE - jQuery Easing
 +
*
 +
* Open source under the BSD License.
 +
*
 +
* Copyright © 2008 George McGinley Smith
 +
* All rights reserved.
 +
*
 +
* Redistribution and use in source and binary forms, with or without modification,
 +
* are permitted provided that the following conditions are met:
 +
*
 +
* Redistributions of source code must retain the above copyright notice, this list of
 +
* conditions and the following disclaimer.
 +
* Redistributions in binary form must reproduce the above copyright notice, this list
 +
* of conditions and the following disclaimer in the documentation and/or other materials
 +
* provided with the distribution.
 +
*
 +
* Neither the name of the author nor the names of contributors may be used to endorse
 +
* or promote products derived from this software without specific prior written permission.
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*
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* MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE ARE DISCLAIMED. IN NO EVENT SHALL THE
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*  COPYRIGHT OWNER OR CONTRIBUTORS BE LIABLE FOR ANY DIRECT, INDIRECT, INCIDENTAL, SPECIAL,
 +
*  EXEMPLARY, OR CONSEQUENTIAL DAMAGES (INCLUDING, BUT NOT LIMITED TO, PROCUREMENT OF SUBSTITUTE
 +
*  GOODS OR SERVICES; LOSS OF USE, DATA, OR PROFITS; OR BUSINESS INTERRUPTION) HOWEVER CAUSED
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* AND ON ANY THEORY OF LIABILITY, WHETHER IN CONTRACT, STRICT LIABILITY, OR TORT (INCLUDING
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*  NEGLIGENCE OR OTHERWISE) ARISING IN ANY WAY OUT OF THE USE OF THIS SOFTWARE, EVEN IF ADVISED
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/*
 +
*
 +
* TERMS OF USE - EASING EQUATIONS
 +
*
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* Open source under the BSD License.
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*
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* Copyright © 2001 Robert Penner
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* All rights reserved.
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*
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* Redistribution and use in source and binary forms, with or without modification,
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* MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE ARE DISCLAIMED. IN NO EVENT SHALL THE
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*  COPYRIGHT OWNER OR CONTRIBUTORS BE LIABLE FOR ANY DIRECT, INDIRECT, INCIDENTAL, SPECIAL,
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*  EXEMPLARY, OR CONSEQUENTIAL DAMAGES (INCLUDING, BUT NOT LIMITED TO, PROCUREMENT OF SUBSTITUTE
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*  GOODS OR SERVICES; LOSS OF USE, DATA, OR PROFITS; OR BUSINESS INTERRUPTION) HOWEVER CAUSED
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*  NEGLIGENCE OR OTHERWISE) ARISING IN ANY WAY OUT OF THE USE OF THIS SOFTWARE, EVEN IF ADVISED
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* OF THE POSSIBILITY OF SUCH DAMAGE.
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</script>
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// dark
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$('.timelineLight').timeline({
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openTriggerClass : '.read_more',
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                     <li><a href="https://2017.igem.org/Team:BGIC-Union/Team">members</a></li>
 
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+
 
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                             <ul class="dropdown-menu" >
 
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<li><a href="https://2017.igem.org/Team:BGIC-Union/Demonstrate">demonstrate</a></li>
 
<li><a href="https://2017.igem.org/Team:BGIC-Union/Demonstrate">demonstrate</a></li>
 
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<li><a href="https://2017.igem.org/Team:BGIC-Union/References">references</a></li>
 
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+
                             
 
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     </div>
 
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−
</nav>           <!-- PAGE CONTENT -->
+
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                    <!-- +++++ Projects Section +++++ -->
+
 
 
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<embed src="https://static.igem.org/mediawiki/2017/4/4b/Finalmodeling.pdf" width="800px" height="600px"/>
+
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</div><!-- /row -->
+
<h3>Introduction</h3> </br>
−
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+
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+
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                <!-- END OUR CLIENTS -->
+
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            </div>
+
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        </div>
+
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        <!-- END PAGE CONTENT -->
+
  
−
</div>
+
In the modeling part, we decide to prove that using NASBA to amplify the circulating tumor DNA is necessary and to estimate the optimal concentration of the circulating tumor DNA for detection.  </br>
  
−
 
+
<h3>The influence of cell free DNA on the effectiveness of the detection device: </h3>  </br>
 +
 
 +
In real physiological environment of the plasma, there are many factors influencing the effectiveness of dcas9 protein binding with the targeted DNA sequence (EML4-ALK), the most prominent one of which is cell free DNA. Cell free DNAs come from apoptosis and necrosis of normal cells and may include gene EML4 and gene ALK, which are potential ligands bind with dcas9 protein and lead to a decrease of the effective concentration of NT7-dcas9 and CT7-dcas9 protein. The effective concentration is referred as the concentration of NT7-dcas9 and CT7-dcas9 which bind to the target sequence (EML4-ALK fusion gene), leading to one DNA molecule bears both NT7-dcas9 and CT7-dcas9 to enable the transcription of report gene. In order to determine the magnitude of the influence from the cell free DNAs on the effectiveness of split-T7-dcas9 in binding with EML4-ALK, the following model is made. Assumption that NT7-dcas9 binds with sequence of EML4 and CT7-dcas9 binds with sequence of ALK is made in the formula. 
 +
</br><img src=" https://static.igem.org/mediawiki/2017/d/d9/Modelbgic.png" width="800px" class="center" /></br>
 +
As shown in the formula above, given the total concentration of NT7-dcas9 and CT7-dcas9, the effective concentration of NT7-dcas9 and CT7-dcas9 decreases as the concentration of cell free DNA increase. In real physiological environment of plasma, the concentration of cell free DNAs is seven orders of magnitude larger than the concentration of circulating tumor DNAs. Therefore, it can be assumed that d[EML4] and d[ALK] approach to infinity. Hence, in this scenario, the change in effective concentration of NT7-dcas9 and CT7-dcas9 is equal to negative infinity, revealed as no signal emitted even if EML4-ALK fusion gene exists.
 +
Based on the modeling, in order to gain valid result of the presence and concentration of EML4-ALK fusion gene, amplifying the concentration of EML4-ALK fusion gene is necessary. By employing the Nucleic Acid Sequence-Based Amplification (NASBA) <strong>(Figure 1)</strong>, the concentration of EML4-ALK fusion gene increases at an exponential rate<strong> (Figure 2) </strong>. Within eight rounds, the concentration of EML4-ALK approaches to a detectable value. Therefore, it is proved that NASBA is a necessary and effective signal amplification approach before detection. 
 +
 
 +
</br><img src=" https://static.igem.org/mediawiki/2017/2/2d/Model21150.png" width="800px" class="center" /> </br> </br><img src=" https://static.igem.org/mediawiki/2017/2/2d/11503model.png" width="800px" class="center" />
 +
 
 +
 +
</br>
 +
<h3>Determining the upper limit of target DNA concentration: </h3></br>
 +
We decide to determine whether there is an upper limit of the target DNA concentration for split-T7-dcas9 system, since it is hypothesized that the effective concentration of NT7-dcas9 and CT7-dcas9 will decrease after the concentration of the target DNA reaches an upper limit; since the increase in potential binding sites increases the probability of one DNA molecule bears only one protein (Figure 3). The upper limit is the optimal point for the highest activity of the detection device. </br>
 +
<img src=" https://static.igem.org/mediawiki/2017/2/2a/11504model.png" width="800px" class="center" /></br></br>
 +
 
 +
To determine the optimal point for detection, the concentration of NT7-dcas9, the concentration of CT7-dcas9, and their fraction of binding with DNA are considered. The formula for the optimal point is established as: </br>
 +
<div class="center">2 × [DNA<sub>optimal</sub>] = [NT7 total] × θN + [CT7 total] × θC, </div></br>
 +
 
 +
where [DNAoptimal] represents the optimal concentration of DNA for detection system. 
 +
In the situation that the concentration of DNA is lower than the optimal concentration, the concentration of NT7-dcas9 and CT7-dcas9 binding with the DNA increases linearly as the concentration of target DNA increases. The change in effective concentration of NT7-dcas9 and CT7-dcas9 are represented in the formula below: </br>
 
    
 
    
−
  </body>
+
<img src=" https://static.igem.org/mediawiki/2017/e/e3/Model5.png" width="800px" class="center" /></br></br>
  
 +
 
 +
 +
</br>When the concentration of target DNA is really low, the dissociation between the DNA and the proteins almost does not occur, therefore, the dissociation constant is negligible. However, after the concentration of DNA reaches a tipping point, the dissociation becomes prominent, thus, θ is considered and the value of the slope decreases by θ.  </br>
  
 +
After the concentration reaches the optimal concentration for detection, the probability of one DNA molecule bearing both NT7-dcas9 and CT7-dcas9 and enabling transcription of report gene decreases as the concentration continually increases. The probability of one DNA molecule bearing both NT7-dcas9 and CT7dcas9 is represented as the formula below: </br></br>
  
−
     
+
 
 +
 
 +
<img src=" https://static.igem.org/mediawiki/2017/d/d9/Modelbgic.png" width="800px" class="center" /></br></br>
 +
 
 +
</br>
 +
Employing the formula above and the data from Peking 2015, the best-fitted curve for the relationship between relative fluorescent unit and target DNA concentration before and after the optimal concentration is reached has been made in<strong> Figure 4</strong> and <strong>Figure 5</strong>. Revealed in the curves, before the optimal concentration is reaches, in the interval of low concentration (0-0.6), the line has a greater slope than the slope after the concentration reaches a certain value (0.6-1.5), decreasing by multiplying the factor of θ. After the optimal concentration is reaches, the slope is negative and the value of the negativity increases as a factor of 公示图 . </br>
 +
 
 +
Therefore, the best-fitted curves prove that our hypothesis is correct.</br>
 +
<img src=" https://static.igem.org/mediawiki/2017/4/44/Model6.png" width="800px" class="center" /></br></br>
 +
<img src=" https://static.igem.org/mediawiki/2017/c/c6/Model7.png" width="800px" class="center" /></br></br>
 +
 
 +
<h3>Conclusion: </h3></br>
 +
From the two models made above, we conclude that using NASBA to amplify the target DNA (EML4-ALK) is necessary and efficient for the detection device in order to avoid the interference of cell free DNAs from normal cells. Also, there is an upper limit of target DNA concentration for the detection device to create the most intense signal. The upper limit of the concentration of DNA is determined by the concentration of NT7-dcas9, CT7-dcas9, and their dissociation constant with target DNA. </br>
 +
</br>
 +
 
 +
 +
<h3>Reference: </h3></br>
 +
 
 +
Zhang Y, Qian L, Wei W, et al. Paired Design of dCas9 as a Systematic Platform for the Detection of Featured Nucleic Acid Sequences in Pathogenic Strains.[J]. Acs Synthetic Biology, 2017, 6(2):211. </br>
 +
1.
 +
2. Pardee K, Green A A, Takahashi M K, et al. Rapid, Low-Cost Detection of Zika Virus Using Programmable Biomolecular
 +
Components[J]. Cell, 2016, 165(5):1255. </br>
 +
 
 +
Han T, Chen Q, Liu H. Engineered photoactivatable genetic switches based on the bacterium phage T7 RNA polymerase[J]. Acs Synthetic Biology, 2016, 6(2). </br>
 +
3.
 +
 
 +
</div>
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 +
 +
 +
 
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  </body>
  
 
</html>
 
</html>

Revision as of 02:21, 2 November 2017

Team：BGIC-Union/Practices-Design

Introduction


In the modeling part, we decide to prove that using NASBA to amplify the circulating tumor DNA is necessary and to estimate the optimal concentration of the circulating tumor DNA for detection.

The influence of cell free DNA on the effectiveness of the detection device:


In real physiological environment of the plasma, there are many factors influencing the effectiveness of dcas9 protein binding with the targeted DNA sequence (EML4-ALK), the most prominent one of which is cell free DNA. Cell free DNAs come from apoptosis and necrosis of normal cells and may include gene EML4 and gene ALK, which are potential ligands bind with dcas9 protein and lead to a decrease of the effective concentration of NT7-dcas9 and CT7-dcas9 protein. The effective concentration is referred as the concentration of NT7-dcas9 and CT7-dcas9 which bind to the target sequence (EML4-ALK fusion gene), leading to one DNA molecule bears both NT7-dcas9 and CT7-dcas9 to enable the transcription of report gene. In order to determine the magnitude of the influence from the cell free DNAs on the effectiveness of split-T7-dcas9 in binding with EML4-ALK, the following model is made. Assumption that NT7-dcas9 binds with sequence of EML4 and CT7-dcas9 binds with sequence of ALK is made in the formula.

As shown in the formula above, given the total concentration of NT7-dcas9 and CT7-dcas9, the effective concentration of NT7-dcas9 and CT7-dcas9 decreases as the concentration of cell free DNA increase. In real physiological environment of plasma, the concentration of cell free DNAs is seven orders of magnitude larger than the concentration of circulating tumor DNAs. Therefore, it can be assumed that d[EML4] and d[ALK] approach to infinity. Hence, in this scenario, the change in effective concentration of NT7-dcas9 and CT7-dcas9 is equal to negative infinity, revealed as no signal emitted even if EML4-ALK fusion gene exists. Based on the modeling, in order to gain valid result of the presence and concentration of EML4-ALK fusion gene, amplifying the concentration of EML4-ALK fusion gene is necessary. By employing the Nucleic Acid Sequence-Based Amplification (NASBA) (Figure 1), the concentration of EML4-ALK fusion gene increases at an exponential rate (Figure 2) . Within eight rounds, the concentration of EML4-ALK approaches to a detectable value. Therefore, it is proved that NASBA is a necessary and effective signal amplification approach before detection.



Determining the upper limit of target DNA concentration:


We decide to determine whether there is an upper limit of the target DNA concentration for split-T7-dcas9 system, since it is hypothesized that the effective concentration of NT7-dcas9 and CT7-dcas9 will decrease after the concentration of the target DNA reaches an upper limit; since the increase in potential binding sites increases the probability of one DNA molecule bears only one protein (Figure 3). The upper limit is the optimal point for the highest activity of the detection device.


To determine the optimal point for detection, the concentration of NT7-dcas9, the concentration of CT7-dcas9, and their fraction of binding with DNA are considered. The formula for the optimal point is established as:
2 × [DNAoptimal] = [NT7 total] × θN + [CT7 total] × θC,

where [DNAoptimal] represents the optimal concentration of DNA for detection system. In the situation that the concentration of DNA is lower than the optimal concentration, the concentration of NT7-dcas9 and CT7-dcas9 binding with the DNA increases linearly as the concentration of target DNA increases. The change in effective concentration of NT7-dcas9 and CT7-dcas9 are represented in the formula below:



When the concentration of target DNA is really low, the dissociation between the DNA and the proteins almost does not occur, therefore, the dissociation constant is negligible. However, after the concentration of DNA reaches a tipping point, the dissociation becomes prominent, thus, θ is considered and the value of the slope decreases by θ.
After the concentration reaches the optimal concentration for detection, the probability of one DNA molecule bearing both NT7-dcas9 and CT7-dcas9 and enabling transcription of report gene decreases as the concentration continually increases. The probability of one DNA molecule bearing both NT7-dcas9 and CT7dcas9 is represented as the formula below:




Employing the formula above and the data from Peking 2015, the best-fitted curve for the relationship between relative fluorescent unit and target DNA concentration before and after the optimal concentration is reached has been made in Figure 4 and Figure 5. Revealed in the curves, before the optimal concentration is reaches, in the interval of low concentration (0-0.6), the line has a greater slope than the slope after the concentration reaches a certain value (0.6-1.5), decreasing by multiplying the factor of θ. After the optimal concentration is reaches, the slope is negative and the value of the negativity increases as a factor of 公示图 .
Therefore, the best-fitted curves prove that our hypothesis is correct.




Conclusion:


From the two models made above, we conclude that using NASBA to amplify the target DNA (EML4-ALK) is necessary and efficient for the detection device in order to avoid the interference of cell free DNAs from normal cells. Also, there is an upper limit of target DNA concentration for the detection device to create the most intense signal. The upper limit of the concentration of DNA is determined by the concentration of NT7-dcas9, CT7-dcas9, and their dissociation constant with target DNA.

Reference:


Zhang Y, Qian L, Wei W, et al. Paired Design of dCas9 as a Systematic Platform for the Detection of Featured Nucleic Acid Sequences in Pathogenic Strains.[J]. Acs Synthetic Biology, 2017, 6(2):211.
1. 2. Pardee K, Green A A, Takahashi M K, et al. Rapid, Low-Cost Detection of Zika Virus Using Programmable Biomolecular Components[J]. Cell, 2016, 165(5):1255.
Han T, Chen Q, Liu H. Engineered photoactivatable genetic switches based on the bacterium phage T7 RNA polymerase[J]. Acs Synthetic Biology, 2016, 6(2).
3.