Difference between revisions of "Team:ZJU-China/Newparts"
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| − | <title> | + | <title>New Parts</title> |
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| − | <link rel="stylesheet" type="text/css" href="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/CSS/barcss/site&action=raw&ctype=text/css" /> | + | <link rel="stylesheet" type="text/css" href="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/CSS/barcss/site.css&action=raw&ctype=text/css" /> |
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<script type="text/javascript" src="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/JS/barjs/main&action=raw&ctype=text/javascript"></script> | <script type="text/javascript" src="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/JS/barjs/main&action=raw&ctype=text/javascript"></script> | ||
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</head> | </head> | ||
| − | <body style="background-image: url('https://static.igem.org/mediawiki/2020/ | + | <body style="background-image: url('https://static.igem.org/mediawiki/2020/8/89/T--ZJU-China--wiki_index_backpage.png');"> |
| − | <!-- Pre Loader Area start --> | + | <!-- Pre Loader Area start --> |
| − | <div id="preloader | + | <div id="preloader" style="text-align:center;margin: 0 auto;"> |
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| − | </ul> | + | </div> |
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| + | <!-- Pre Loader Area End --> | ||
| + | |||
| + | <div class="navigation"> | ||
| + | |||
| + | <ul> | ||
| + | <li> | ||
| + | <a href="#Cloning" style="background-color: #f6b37f;" class="nav-active rounded"><span>Overview</span></a> | ||
| + | </li> | ||
| + | <li> | ||
| + | <a href="#about" style="background-color: #7ecef4;" class="rounded"><span>Improved T7 Promoters</span></a> | ||
| + | </li> | ||
| + | <li> | ||
| + | <a href="#services" style="background-color: #f19ec2;" class="rounded"><span><i>E.coli</i> BL21 (DE3)</span></a> | ||
| + | </li> | ||
| + | <li> | ||
| + | <a href="#showcase" style="background-color:#eb6877;"class="rounded"><span><i>E.coli</i> SHuffle<sup>®</sup> T7 Express Cells</span></a> | ||
| + | </li> | ||
| + | |||
| + | |||
| + | |||
| + | </ul> | ||
| + | </div> | ||
| + | |||
<!--Main Menu/ Mobile Menu Section--> | <!--Main Menu/ Mobile Menu Section--> | ||
<section class="menu-section-area" > | <section class="menu-section-area" > | ||
<!-- Navigation --> | <!-- Navigation --> | ||
| − | <nav class="navbar navbar-expand-lg fixed-top d-none d-sm-none d-md-block d-lg-block d-xl-block" id="mainNav" style="background- | + | <nav class="navbar navbar-expand-lg fixed-top d-none d-sm-none d-md-block d-lg-block d-xl-block" id="mainNav" style="background-color:white;" > |
<div class="container"> | <div class="container"> | ||
| − | <a class="navbar-brand" href=" | + | <a class="navbar-brand" href="https://2020.igem.org/Team:ZJU-China" style="left:-10%;margin-top:-18px;margin-bottom:-26px;margin-left:-18px;margin-right:-18px;"><img src="https://static.igem.org/mediawiki/2020/d/dd/T--ZJU-China--wiki_logo_new.png" class="img-fluid" width="150px" ></a> |
| − | + | ||
<div class="collapse navbar-collapse" id="navbarResponsive" > | <div class="collapse navbar-collapse" id="navbarResponsive" > | ||
<ul class="navbar-nav text-capitalize ml-auto"> | <ul class="navbar-nav text-capitalize ml-auto"> | ||
<li class="nav-item dropdown-box" style="background-color:pink;;opacity: 0.9;"> | <li class="nav-item dropdown-box" style="background-color:pink;;opacity: 0.9;"> | ||
| − | <a class="nav-link js-scroll-trigger" href=" | + | <a class="nav-link js-scroll-trigger" href="https://2020.igem.org/Team:ZJU-China">Home</a> |
</li> | </li> | ||
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<a class="nav-link" href="#">Team</a> | <a class="nav-link" href="#">Team</a> | ||
<ul class="dropdown-list" > | <ul class="dropdown-list" > | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Team">Members</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Attributions">Attributions</a></li> |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Medals">Medals</a></li> | ||
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<a class="nav-link" href="#">Project</a> | <a class="nav-link" href="#">Project</a> | ||
<ul class="dropdown-list"> | <ul class="dropdown-list"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Description">Description</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Background">Background</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Design">Design</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Magnetosome">Magnetosome</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Experiments">Experiments</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Results">Results</a></li> |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Future">Future</a></li> | ||
</ul> | </ul> | ||
</li> | </li> | ||
<li class="nav-item dropdown-box" style="background-color:dodgerblue;opacity: 0.9;"> | <li class="nav-item dropdown-box" style="background-color:dodgerblue;opacity: 0.9;"> | ||
| − | <a class="nav-link" href="#"> | + | <a class="nav-link" href="#">Model</a> |
<ul class="dropdown-list"> | <ul class="dropdown-list"> | ||
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Model">Model</a></li> | |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Engineering">Engineering</a></li> |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Proof_Of_Concept">Proof of Concept</a></li> | ||
| + | |||
</ul> | </ul> | ||
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<a class="nav-link" href="#">Parts</a> | <a class="nav-link" href="#">Parts</a> | ||
<ul class="dropdown-list"> | <ul class="dropdown-list"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Contribution">Contribution</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Improve">Improve</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Newparts">New Parts</a></li> |
</ul> | </ul> | ||
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<a class="nav-link" href="#">Human</a> | <a class="nav-link" href="#">Human</a> | ||
<ul class="dropdown-list"> | <ul class="dropdown-list"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Human_Practices">Human Practices</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Education">Education</a></li> |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Collaborations">Collaborations</a></li> | ||
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Partnership">Partnership</a></li> | ||
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Safety">Safety</a></li> | ||
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Implementation">Implementation</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Entrepreneurship">Entrepreneurship</a></li> | |
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| − | <li><a href=" | + | |
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</ul> | </ul> | ||
</li> | </li> | ||
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</ul> | </ul> | ||
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<!-- Mobile Menu Start --> | <!-- Mobile Menu Start --> | ||
<nav class="mobile_menu hidden d-none"> | <nav class="mobile_menu hidden d-none"> | ||
| − | <a href=" | + | <a href="https://2020.igem.org/Team:ZJU-China"><img class="mobile-logo" src="https://static.igem.org/mediawiki/2020/7/7f/T--ZJU-China--temt.png" alt="Logo"></a> |
<ul class="nav navbar-nav navbar-right menu"> | <ul class="nav navbar-nav navbar-right menu"> | ||
<li class="dropdown active"> | <li class="dropdown active"> | ||
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<a>Team</a> | <a>Team</a> | ||
<ul class="sub_menu"> | <ul class="sub_menu"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Team">Members</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Attributions">Attributions</a></li> |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Medals">Medals</a></li> | ||
| + | |||
</ul> | </ul> | ||
</li> | </li> | ||
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<a>Project</a> | <a>Project</a> | ||
<ul class="sub_menu"> | <ul class="sub_menu"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Description">Description</a></li> |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Background">Background</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Design">Design</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Magnetosome">Magnetosome</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Experiments">Experiments</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Results">Results</a></li> | |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Future">Future</a></li> | ||
</ul> | </ul> | ||
</li> | </li> | ||
<li class="dropdown"> | <li class="dropdown"> | ||
| − | <a> | + | <a>Model</a> |
<ul class="sub_menu"> | <ul class="sub_menu"> | ||
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Model">Model</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Engineering">Engineering</a></li> | |
| + | <li><a href="https://2020.igem.org/Team:ZJU-China/Proof_Of_Concept">Proof of Concept</a></li> | ||
| + | |||
</ul> | </ul> | ||
</li> | </li> | ||
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<a>Parts</a> | <a>Parts</a> | ||
<ul class="sub_menu"> | <ul class="sub_menu"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Contribution">Contribution</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Improve">Improve</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Newparts">New Parts</a></li> |
| + | |||
</ul> | </ul> | ||
</li> | </li> | ||
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<a>Human</a> | <a>Human</a> | ||
<ul class="sub_menu"> | <ul class="sub_menu"> | ||
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Human_Practices">Human Practices</a></li> |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Education">Education</a></li> |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Collaborations">Collaborations</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Partnership">Partnership</a></li> | |
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Safety">Safety</a></li> | |
| − | + | ||
| − | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Implementation">Implementation</a></li> | |
| − | <li><a href=" | + | <li><a href="https://2020.igem.org/Team:ZJU-China/Entrepreneurship">Entrepreneurship</a></li> |
| − | <li><a href=" | + | |
</ul> | </ul> | ||
</li> | </li> | ||
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</ul> | </ul> | ||
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<!--Main Menu/ Mobile Menu Section--> | <!--Main Menu/ Mobile Menu Section--> | ||
| − | <div | + | <div id="swrapper"> |
| − | + | <div id="mcontent" > | |
| − | + | ||
| − | + | ||
| − | + | ||
| − | <div id=" | + | |
| + | <div class="pagename"> | ||
| + | <h1>Newparts</h1> | ||
| + | </div> | ||
| − | <div class="pagestyle" style="background-image: url('https://static.igem.org/mediawiki/2020/c/cd/T--ZJU-China--wiki_navback.jpg') | + | |
| − | + | <div id="scrollable"> | |
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| − | + | <div class="pagestyle" style="background-image: url('https://static.igem.org/mediawiki/2020/c/cd/T--ZJU-China--wiki_navback.jpg');"> | |
| − | + | <div class="section Cloning" id="Cloning"> | |
| − | + | <div class="container1"> | |
| − | + | ||
| − | + | <h2>Overview</h2> | |
| − | + | <p style="color:#808080;line-height:200%"><b> | |
| − | + | This year, we designed 5 parts, and all of them had been submitted to the iGEM Registry following the BioBricks assembly standard. | |
| − | + | </p> | |
| − | + | <p> | |
| − | + | ||
| − | + | </p> | |
| − | + | <br> | |
| − | + | <p style="color:#808080;line-height:200%"><b> | |
| − | <div class="imgbox"> | + | Here is a brief description of our parts. For more detailed information about our parts, we strongly recommend you visit the corresponding parts pages on the iGEM BioBricks Registry. |
| − | + | </p> | |
| − | + | ||
| − | + | <br> | |
| − | + | <div class="imgbox"> | |
| − | + | <h6 style="font-size:large;">Parts Table</h6> | |
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| − | < | + | </div> |
| − | <tr> | + | <div class="imgbox"> |
| − | <td>BBa_K3398000</td> | + | <table style="width: auto;height: auto;max-width: 100%;max-height: 100%;margin-right:auto ;margin-left:auto;" class="pure-table pure-table-bordered"> |
| − | + | <thead> | |
| − | + | <tr> | |
| − | <td>64</td> | + | <th>Name</th> |
| − | <td>Mingxiao Wei</td> | + | <th>Type</th> |
| − | + | <th>Description</th> | |
| + | <th>Length(bp)</th> | ||
| + | <th>Designer</th> | ||
| + | </tr> | ||
| + | </thead> | ||
| + | |||
| + | <tbody> | ||
| + | <tr> | ||
| + | <td>BBa_K3398000</td> | ||
| + | <td>T7</td> | ||
| + | <td>Modified T7 promoter 1</td> | ||
| + | <td>64</td> | ||
| + | <td>Mingxiao Wei</td> | ||
| + | </tr> | ||
| + | |||
| + | <tr> | ||
| + | <td>BBa_K3398001</td> | ||
| + | <td>T7</td> | ||
| + | <td>Modified T7 promoter 2</td> | ||
| + | <td>64</td> | ||
| + | <td>Mingxiao Wei</td> | ||
| + | </tr> | ||
| + | |||
| + | <tr> | ||
| + | <td>BBa_K3398002</td> | ||
| + | <td>T7</td> | ||
| + | <td>Modified T7 promoter 3</td> | ||
| + | <td>64</td> | ||
| + | <td>Mingxiao Wei</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>BBa_K3398003</td> | ||
| + | <td>coding</td> | ||
| + | <td>mamC_ZZ fusion protein</td> | ||
| + | <td>771</td> | ||
| + | <td>Mingxiao Wei</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>BBa_K3398004</td> | ||
| + | <td>coding</td> | ||
| + | <td>mamC_ZZ fusion protein with GST tag</td> | ||
| + | <td>1464</td> | ||
| + | <td>Mingxiao Wei</td> | ||
| + | </tr> | ||
| + | |||
| + | <tr> | ||
| + | <td>BBa_K3398005</td> | ||
| + | <td>coding</td> | ||
| + | <td>scFv_Fc fusion protein</td> | ||
| + | <td>1497</td> | ||
| + | <td>Xiner Ying</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>BBa_K3398006</td> | ||
| + | <td>coding</td> | ||
| + | <td>scFv_Fc protein with FLAG tag</td> | ||
| + | <td>1524</td> | ||
| + | <td>Xiner Ying</td> | ||
| + | </tr> | ||
| + | |||
| + | </tbody> | ||
| + | </table> | ||
| + | </div> | ||
| + | <br> | ||
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</div> | </div> | ||
| − | < | + | </div> |
| + | |||
| + | |||
| + | <div class="section about" id="about"> | ||
| + | <div class="container1"> | ||
| + | <h2>Improved T7 Promoters</h2> | ||
| + | <p> | ||
| + | <a href="http://parts.igem.org/Part:BBa_K3398000" target="_blank" style="color:#0a8aff;">BBa_K3398000</a>, <a href="http://parts.igem.org/Part:BBa_K3398001" target="_blank" style="color:#0a8aff;">BBa_K3398001</a>, <a href="http://parts.igem.org/Part:BBa_K3398002" target="_blank" style="color:#0a8aff;">BBa_K3398002</a> | ||
| + | </p> | ||
| + | |||
| + | <br> | ||
| + | |||
| + | |||
| + | <p> | ||
| + | As a widely used promoter for expression of heterogenous protein in some <i>E.coli</i> strains such as BL21 (DE3) for its high efficiency, we improved T7 promoter by introducing some point mutations distributing in the range from +25 to +31 of the T7 promoter sequence. We chose EGFP as a reporter to test the function of our modified T7 promoters. | ||
| + | </p> | ||
| + | |||
| − | + | </div> | |
| − | + | ||
</div> | </div> | ||
| − | + | <div class="section services" id="services"> | |
| − | + | <div class="container1"> | |
| − | + | <h2 style="line-height:1.5;">Parts expressed by <i>E.coli</i> BL21 (DE3)</span></a> </h2> | |
| − | + | <br> | |
| − | + | ||
| − | + | ||
| − | <h2> | + | |
<p> | <p> | ||
| − | + | <a href="http://parts.igem.org/Part:BBa_K3398003" target="_blank" style="color:#0a8aff;">BBa_K3398003</a>, <a href="http://parts.igem.org/Part:BBa_K3398004" target="_blank" style="color:#0a8aff;">BBa_K3398004</a> | |
</p> | </p> | ||
| − | |||
<br> | <br> | ||
| − | + | <p> | |
| + | These parts are coding sequences of mamC and ZZ fusion protein. And for separation from other proteins at the purification step, we introduced a GST tag at the N-terminus and a thrombin site between the fusion protein and tag for cleavage after purification. To maximize the expression of a functional protein, they were subcloned into gene expression vector pGEX-2TK and transformed into <i>E.coli</i> BL21 (DE3) to express. These parts can also be expressed by magnetotactic bacteria such as MSR-1 to produce magnetosomes modified with ZZ protein on the surface as a self-assembly platform for polymer varied antibodies. | ||
| + | </p> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div class="section showcase" id="showcase"> | ||
| + | <div class="container1"> | ||
| + | <h2 style="line-height:1.5;">Parts expressed by <i>E.coli</i> SHuffle<sup>®</sup> T7 Express Cells</h2> | ||
<p> | <p> | ||
| − | + | <a href="http://parts.igem.org/Part:BBa_K3398005"target="_blank" style="color:#0a8aff;">BBa_K3398005</a>, <a href="http://parts.igem.org/Part:BBa_K3398006"target="_blank" style="color:#0a8aff;">BBa_K3398006</a> | |
</p> | </p> | ||
| − | + | <br> | |
| − | + | <p> | |
| − | + | These parts are coding sequences of scFv and Fc proteins. And for separation from other proteins at the purification step, we introduced a 10× histidine tag at the N-terminus which was followed by an enterokinase cleavage site for cleavage after purification. To maximize the expression of a functional protein, they were subcloned into gene expression vector pET19b and transformed into <i>E.coli</i> BL21 (DE3) to express. As the redox potential in the cytoplasm of general <i>E.coli</i> such as BL21 is too high, the disulfide bond in scFv will be reduced into thiol, which may not express the functional fusion proteins. So we chose <i>E.coli</i> SHuffle<sup>®</sup> with a reduced cytoplasmic environment as the chassis organism to express scFv-Fc fusion protein. The part of scFv-Fc is ideal molecule which can bind HER2 cells strongly and specifically, by adding report genes at either C- or the N-terminus, they can be used for the detection of HER2 cells. | |
| − | + | </p> | |
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| + | <div style="display:inline-block;height:180px;margin-left:25%;padding-top:130px;text-align:left;"> | ||
| + | <a href="http://www.zju.edu.cn/" target="_blank" style="color:red;opacity:0;font-size:15px;z-index: 99999;width:20%">aannnaaaaannnnnnnnnnnnn</a> | ||
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| + | <div style="display:inline-block;height:180px;margin-left:15%;margin-right:10%;padding-top:130px;text-align:center;"> | ||
| + | <a href="mailto:liangqiyu22@zju.edu.cn" target="_blank" style="color:red;opacity:0;font-size:15px;z-index: 99999;width:20%">aannnnnnnnn</a> | ||
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| + | </div> | ||
| + | <script type="text/javascript" src="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/jsjquerymin&action=raw&ctype=text/javascript"></script> | ||
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| − | <script type="text/javascript" src="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/ | + | <script type="text/javascript" src="https://2020.igem.org/wiki/index.php?title=Template:ZJU-China/jscustom1&action=raw&ctype=text/javascript"></script> |
| − | < | + | <script> |
| − | + | $document.ready(function () { | |
| + | $("#HQ_page").attr('id', "); | ||
| + | }); | ||
| − | + | </script> | |
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| − | + | <script> | |
| − | + | var typed = new Typed('#typed', { | |
| − | }); | + | stringsElement: '#typed-strings', |
| + | typeSpeed: 100, | ||
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| + | loop: true, | ||
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| − | </script> | + | <script> |
| + | function topMao(target) { | ||
| + | $('html, body').animate({ scrollTop: $(target).offset().top - 170 }, 500);//130为锚点到距顶部的距离，500为执行时间 | ||
| + | return true; | ||
| + | } | ||
| + | </script> | ||
| + | <script> | ||
| + | var topBegin = $("#accordion").offset().top; //获取导航栏（class='positionMiddleNav'）离视口的高度 | ||
| + | $(window).scroll(function () { //scroll事件 | ||
| + | var win_top = $(this).scrollTop(); //获取滚动条位置 | ||
| + | var _top = $("#accordion").offset().top; //获取当前导航栏距视口高度 | ||
| + | console.log(win_top, 'aa'); | ||
| + | console.log(_top, 'cc'); | ||
| + | if (win_top >= _top - 150) { | ||
| − | + | $("#accordion").css({ position: "fixed", top: "10em" }); | |
| − | + | } | |
| − | + | if (win_top < topBegin) { //因为导航栏距视口高度在定位发生后是变化的 ， | |
| − | + | //所以当导航栏回到原位置时保持先前状态需要将滚动条位置与最先前的导航栏位置进行对比 | |
| − | + | $("#accordion").css({ position: "absolute" }); | |
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| − | < | + | function displaywindowsize() { |
| + | var w = document.documentElement.clientWidth; | ||
| + | var h = document.documentElement.clientHeight; | ||
| + | if (w < 1100 && w > 800) { | ||
| − | + | $("#accordion").css({ left: "2%" }); | |
| − | + | $("#accordion").css({ display: "block" }); | |
| − | + | } | |
| − | + | else if (w < 800) { | |
| − | + | $("#accordion").css({ display: "none" }); | |
| − | $("#accordion").css({left:" | + | } |
| − | + | else { | |
| + | $("#accordion").css({ left: "10%" }); | ||
| + | $("#accordion").css({ display: "block" }); | ||
| + | } | ||
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| − | + | window.addEventListener("resize", displaywindowsize); | |
| − | + | displaywindowsize(); | |
| − | </script> | + | </script> |
Latest revision as of 04:00, 28 October 2020
Newparts
Overview
This year, we designed 5 parts, and all of them had been submitted to the iGEM Registry following the BioBricks assembly standard.
Here is a brief description of our parts. For more detailed information about our parts, we strongly recommend you visit the corresponding parts pages on the iGEM BioBricks Registry.
Parts Table
| Name | Type | Description | Length(bp) | Designer |
|---|---|---|---|---|
| BBa_K3398000 | T7 | Modified T7 promoter 1 | 64 | Mingxiao Wei |
| BBa_K3398001 | T7 | Modified T7 promoter 2 | 64 | Mingxiao Wei |
| BBa_K3398002 | T7 | Modified T7 promoter 3 | 64 | Mingxiao Wei |
| BBa_K3398003 | coding | mamC_ZZ fusion protein | 771 | Mingxiao Wei |
| BBa_K3398004 | coding | mamC_ZZ fusion protein with GST tag | 1464 | Mingxiao Wei |
| BBa_K3398005 | coding | scFv_Fc fusion protein | 1497 | Xiner Ying |
| BBa_K3398006 | coding | scFv_Fc protein with FLAG tag | 1524 | Xiner Ying |
Improved T7 Promoters
BBa_K3398000, BBa_K3398001, BBa_K3398002
As a widely used promoter for expression of heterogenous protein in some E.coli strains such as BL21 (DE3) for its high efficiency, we improved T7 promoter by introducing some point mutations distributing in the range from +25 to +31 of the T7 promoter sequence. We chose EGFP as a reporter to test the function of our modified T7 promoters.
Parts expressed by E.coli BL21 (DE3)
These parts are coding sequences of mamC and ZZ fusion protein. And for separation from other proteins at the purification step, we introduced a GST tag at the N-terminus and a thrombin site between the fusion protein and tag for cleavage after purification. To maximize the expression of a functional protein, they were subcloned into gene expression vector pGEX-2TK and transformed into E.coli BL21 (DE3) to express. These parts can also be expressed by magnetotactic bacteria such as MSR-1 to produce magnetosomes modified with ZZ protein on the surface as a self-assembly platform for polymer varied antibodies.
Parts expressed by E.coli SHuffle® T7 Express Cells
These parts are coding sequences of scFv and Fc proteins. And for separation from other proteins at the purification step, we introduced a 10× histidine tag at the N-terminus which was followed by an enterokinase cleavage site for cleavage after purification. To maximize the expression of a functional protein, they were subcloned into gene expression vector pET19b and transformed into E.coli BL21 (DE3) to express. As the redox potential in the cytoplasm of general E.coli such as BL21 is too high, the disulfide bond in scFv will be reduced into thiol, which may not express the functional fusion proteins. So we chose E.coli SHuffle® with a reduced cytoplasmic environment as the chassis organism to express scFv-Fc fusion protein. The part of scFv-Fc is ideal molecule which can bind HER2 cells strongly and specifically, by adding report genes at either C- or the N-terminus, they can be used for the detection of HER2 cells.
