Difference between revisions of "Team:RDFZ-China/Measurement"

 
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                    <ul>
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                        <li><a href="#section1">Expression Burden</a></li>
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                        <li><a href="#section2">Capacity Monitor</a></li>
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                        <li><a href="#section3">VioABDE</a></li>
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                        <li><a href="#section4">Alternative Device</a></li>
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                        <li><a href="#section5">References</a></li>
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        <div class="description">
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            <h1>Measurement</h1>
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            <div class="topic-title" id="section1">
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                <h3>Expression Burden</h3>
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                <p>In synthetic biology, a common issue in integrating exogenous sequences into host cells is that heterologous gene expression can cause significant metabolic burden. Typically, this gene expression burden is implied by a declining growth rate of bacteria, which is unfavorable for fermentation industry because it lowers yields and profits. According to Ceroni et al. (2015), a fluorescence-based method allows to measure gene expression capacity of cells, by inserting a “capacity monitor”.</p>
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            <div class="topic-title" id="section2">
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                <h3>Capacity Monitor</h3>
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                <p>The “Capacity Monitor” consists of a superfold GFP with a strong constitutive promoter, which is constantly expressed in the bacterial genome. Thus, any alternation in fluorescence intensity can reflect in overall resource availability in the cell. Particularly, with expression of synthesized genes, the expression of GFP will decrease, thus reporting a reallocation of cellular resources. </p>
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            <div class="topic-title" id="section3">
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                <h3>VioABDE</h3>
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                <p>After applying capacity monitor, we compared the fluorescence of E. coli with an empty vector and E. coli with VioABDE inserted and expressed. Both strains were incubated for 24 hours with fluorescence and OD constantly monitored. However, the OD of bacteria with VioABDE increased faster than the bacteria without Vio expression, which contradicts our initial expectation.</p>
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                <p>We co transformed the TlpA39-VioABDE into the capcatiy monitor component cell which was made by beyotime component cell kit. </p>
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                <p>Then the Capacity Monitor and Capacity Monitor-Vio were incubicated in microshaker in 39 degree Celsius, then they were added to the 96 well plates.
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                <img src="https://static.igem.org/mediawiki/2018/3/32/T--RDFZ-China--CPVIO.png",width=100%>
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                <img src="https://static.igem.org/mediawiki/2018/2/20/T--RDFZ-China--CM%2BVio.png",width=100% />"
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            </div>
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            <div class="topic-title" id="section4">
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                <h3>Alternative Device</h3>
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                <p>Since we target to modify bacteria used in fermentation industry, which primarily are used for material synthesis and production, our sequence inserts should occupy as limited resources in bacteria as possible to avoid significant expression burden. In order to reduce this expression stress, we designed another device for fermentation which used a LuxR repressive promoter (Peking-S, 2011) and the “Cold Box” in 5’UTR region of CspA (Ionis Paris, 2017). With only one transcriptional regulator, less energy will be consumed, which is beneficial for fermentation industry.</p>
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        <div class="reference">
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            <div class="topic-title" id="section5">
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                <h3>References</h3>
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                <p>Ceroni, F., Algar, R., Stan, G. B., & Ellis, T. (2015). Quantifying cellular capacity identifies gene expression designs with reduced burden. Nature methods, 12(5), 415.</p>
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                <p>Ionis Paris. (2017). Biological thermo-responsive solution, that reduces the plant’s thermal stresses. Available from: https://2017.igem.org/Team:IONIS-PARIS.</p>
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                <p>Peking-S iGEM. (2011). A Chemical Wire Toolbox for Synthetic Microbial Consortia. Avaible from: https://2011.igem.org/Team:Peking_S.</p>
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  .s-section,.css-1c86csr
 
  .s-contact-info-form .s-contact-info-item,.css-1c86csr
 
  .s-blog .s-blog-details-blurb,.css-1c86csr
 
  .s-footer-section .s-footer-text,.css-1c86csr
 
  .s-item-text{font-size:15px;}.css-1c86csr .navbar-drawer-bar:not(.drawer-open) .navbar{background:white;border-radius:4px;}.css-1c86csr .navbar-drawer-bar:not(.drawer-open) .navbar .navbar-icons span{background-color:#666;}.css-1c86csr .s-title{color:#555;}.css-1c86csr .s-subtitle{color:#f50308;}.css-1c86csr .s-item-title,.css-1c86csr
 
  .s-blog-title,.css-1c86csr
 
  .s-ecommerce-card-view-card-name{color:#555;}.css-1c86csr .s-item-subtitle,.css-1c86csr
 
  .s-page-product .s-section .s-ecommerce .s-ecommerce-row-view-product .s-ecommerce-row-view-product-detail-panel .s-ecommerce-row-view-product-pricing,.css-1c86csr
 
  .slides .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-cards .s-ecommerce-card-view-card .s-ecommerce-card-view-card-price,.css-1c86csr
 
  .s-blog-info{color:#9b59b6;}.css-1c86csr .s-item-text,.css-1c86csr
 
  .s-footer-text,.css-1c86csr
 
  .s-blog-details-blurb{color:#50555c;}.css-1nxcohz .s-section:not(.s-slider-section):not(.s-grid-section),
 
    .css-1nxcohz .s-grid-section:not(._wide){padding-top:75px;padding-bottom:75px !important;-webkit-box-sizing:border-box;box-sizing:border-box;min-height:calc(100vh - 0px);}.css-10c7nxj .s-section:not(.s-slider-section):not(.s-grid-section),
 
    .css-10c7nxj .s-grid-section:not(._wide){padding-top:calc(0px + 75px);padding-bottom:75px ;}</style><div data-hypernova-key="SiteBootstrapper" data-hypernova-id="a4d5cd86-af29-4d11-8a6f-302d6f8e007b" data-hypernova-emotion-id="enscdp,1ym1w9s,3109jj,1c86csr,1nxcohz,10c7nxj"><div data-reactroot=""><div id="s-content" class="s-variation-default s-custom-colors s-animation-image_link_hover-dark_overlay s-font-heading-source-sans-pro s-font-title-source-sans-pro s-font-body-nunito s-entrance-mask s-theme-alignment-center" lang="zh-CN"><div style="margin:auto"><div class="social-thumbnail"><div style="display:none"><img src="https://static-assets.strikinglycdn.com/images/fb_images/default.png" width="300" height="300"/></div></div><div><style id="color-style-tag">.s-custom-colors .s-bg-overlay:before{background:rgba(126,8,8,0.35);}.s-custom-colors .s-component.s-social-media ul.s-social-media-buttons .s-social-icon{background:rgba(155,89,182,0.5);}.s-custom-colors .s-ecommerce-row-view-product .s-ecommerce-row-view-product-thumbnail-list ul li.current{border: 2px solid #9b59b6}.s-custom-colors .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-card .s-ecommerce-card-view-card-stock-warning{background: #9b59b6;}.s-custom-colors .s-ecommerce-shopping-cart-wrapper .s-ecommerce-shopping-cart{background: #9b59b6;}.s-custom-colors .s-ecommerce-shopping-cart-wrapper .s-ecommerce-shopping-cart.hovered{background: #9b59b6;}.s-custom-colors .s-ecommerce-shopping-cart-wrapper .s-ecommerce-shopping-cart-mobile{
 
        background: #fed5d5;
 
        border-top: 1px solid #cbaaaa;}.s-custom-colors .s-ecommerce-shopping-cart-wrapper .s-ecommerce-shopping-cart-mobile .check-btn{background: #9b59b6;}.s-custom-colors .s-ecommerce-animation-item{
 
        border: 2px solid #9b59b6;
 
        color: #9b59b6;}.s-custom-colors .s-ecommerce-row-view-product .mobile-select .price-label{color: #9b59b6}.s-custom-colors .s-ecommerce-row-view-product .mobile-select .variation-item.selected{border: 1px solid #9b59b6; background: #9b59b6;}.s-custom-colors .s-ecommerce-row-view-product .mobile-select .add-btn{background: #9b59b6;}.s-custom-colors .s-ecommerce-row-view-product .slider-wrapper .slider-dot-wrapper .slider-dot{border: 1px solid #9b59b6}.s-custom-colors .s-ecommerce-row-view-product .slider-wrapper .slider-dot-wrapper .slider-dot.selected{background: #9b59b6}.s-custom-colors .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-cards .s-ecommerce-card-view-card .s-ecommerce-card-view-card-price{color: #9b59b6;}.s-custom-colors .s-ecommerce .s-ecommerce-row-view-product .s-ecommerce-row-view-product-detail-panel .s-ecommerce-row-view-product-pricing{color: #9b59b6;}.s-custom-colors .s-section .s-category-bar .category-list .category-link-item.selected{color: #9b59b6;}.s-custom-colors .s-section .s-category-bar .category-list .category-link-item.selected:after{background: #9b59b6}.s-custom-colors .s-section .s-category-bar .category-list .category-link-item-wrapper:hover .category-link-item:not(.no-hover){color: #9b59b6;}.s-custom-colors .s-ecommerce .s-ecommerce-products-wrapper .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-detail .s-ecommerce-card-view-detail-header .next-product-btn{color: #9b59b6;}.s-custom-colors .s-ecommerce .s-ecommerce-products-wrapper .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-detail .s-ecommerce-card-view-detail-header .prev-product-btn{color: #9b59b6;}.s-custom-colors .s-ecommerce .s-ecommerce-products-wrapper .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-detail .s-ecommerce-card-view-detail-header .back-btn{color: #9b59b6;}.s-custom-colors .s-ecommerce .s-ecommerce-products-wrapper .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-detail .s-ecommerce-card-view-detail-header .next-product-btn:hover{color: #af7ac4;}.s-custom-colors .s-ecommerce .s-ecommerce-products-wrapper .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-detail .s-ecommerce-card-view-detail-header .prev-product-btn:hover{color: #af7ac4;}.s-custom-colors .s-ecommerce .s-ecommerce-products-wrapper .s-ecommerce-card-view-wrapper .s-ecommerce-card-view-detail .s-ecommerce-card-view-detail-header .back-btn:hover{color: #af7ac4;}.s-custom-colors .s-donation-section .s-component.s-donation .s-donation-progress .progress-bar .progress{background: #9b59b6}.s-custom-colors .s-donation-section .s-component.s-donation .s-donation-progress .progress-bar .indicator:before{background: #9b59b6}.s-custom-colors .s-donation-section .s-component.s-donation .s-donation-progress .progress-bar .indicator:after{border-top-color: #9b59b6}
 
.s-custom-colors .navigator .nav-container .nav ul li a:hover,
 
.s-custom-colors .navigator .nav-container .nav ul li a.selected {
 
  background: #feeaea;
 
}
 
.s-custom-colors .navigator .nav-container .nav ul.items li,
 
.s-custom-colors .navigator .nav-container .nav ul.items,
 
.s-custom-colors .navigator {
 
  border-color: #fec0c0;
 
}
 
 
.s-custom-colors .dummy-shadow {
 
  box-shadow: #fed5d5 0 0 10px 0 inset;
 
}
 
.navbar-drawer.strikingly-drawer ul#nav-drawer-list {
 
  border-right: 1px solid #feeaea;
 
}
 
.navbar-drawer.strikingly-drawer ul#nav-drawer-list li a.selected {
 
  background: #feeaea;
 
  box-shadow: 5px 0 0 0 #fec0c0 inset;
 
}
 
.navbar-drawer.strikingly-drawer ul#nav-drawer-list li a {
 
  border-bottom: 1px solid #fec0c0;
 
}.s-custom-colors .s-text-color-custom1{ color: #9b59b6; }
 
.s-custom-colors .s-text-color-custom2{ color: #fc1010; }
 
</style><style id="font-style-tag">
 
    #s-content.s-font-body-nunito .s-font-body {
 
      font-family: nunito, helvetica, sans-serif;
 
    }
 
    #s-content.s-font-body-nunito .s-font-body:lang(ja) {
 
      font-family: nunito, helvetica, "ヒラギノ角ゴ Pro W3","Hiragino Kaku Gothic Pro",Osaka,"メイリオ",Meiryo,"MS Pゴシック","MS PGothic", sans-serif
 
    }
 
    #s-content.s-font-body-nunito .s-font-body:lang(zh-cn),
 
    #s-content.s-font-body-nunito .s-font-body:lang(sxl),
 
    #s-content.s-font-body-nunito .s-font-body:lang(zh) {
 
      font-family: nunito, helvetica, 'PingFang SC',"Microsoft YaHei","微软雅黑",STXihei,"华文细黑", sans-serif
 
    }
 
    #s-content.s-font-body-nunito .s-font-body:lang(zh-tw) {
 
      font-family: nunito, helvetica, 'Lisong Pro', 'PMingLiU', 'PingFang TC','Microsoft JhengHei',"微軟正黑體",STXihei, sans-serif;
 
    }
 
 
 
    #s-content.s-font-title-source-sans-pro .s-font-title {
 
      font-family: "source sans pro", helvetica, sans-serif;
 
    }
 
    #s-content.s-font-title-source-sans-pro .s-font-title:lang(ja) {
 
      font-family: "source sans pro", helvetica, "ヒラギノ角ゴ Pro W3","Hiragino Kaku Gothic Pro",Osaka,"メイリオ",Meiryo,"MS Pゴシック","MS PGothic", sans-serif
 
    }
 
    #s-content.s-font-title-source-sans-pro .s-font-title:lang(zh-cn),
 
    #s-content.s-font-title-source-sans-pro .s-font-title:lang(sxl),
 
    #s-content.s-font-title-source-sans-pro .s-font-title:lang(zh) {
 
      font-family: "source sans pro", helvetica, 'PingFang SC',"Microsoft YaHei","微软雅黑",STXihei,"华文细黑", sans-serif
 
    }
 
    #s-content.s-font-title-source-sans-pro .s-font-title:lang(zh-tw) {
 
      font-family: "source sans pro", helvetica, 'Lisong Pro', 'PMingLiU', 'PingFang TC','Microsoft JhengHei',"微軟正黑體",STXihei, sans-serif;
 
    }
 
 
 
    #s-content.s-font-heading-source-sans-pro .s-font-heading {
 
      font-family: "source sans pro", helvetica, sans-serif;
 
    }
 
    #s-content.s-font-heading-source-sans-pro .s-font-heading:lang(ja) {
 
      font-family: "source sans pro", helvetica, "ヒラギノ角ゴ Pro W3","Hiragino Kaku Gothic Pro",Osaka,"メイリオ",Meiryo,"MS Pゴシック","MS PGothic", sans-serif
 
    }
 
    #s-content.s-font-heading-source-sans-pro .s-font-heading:lang(zh-cn),
 
    #s-content.s-font-heading-source-sans-pro .s-font-heading:lang(sxl),
 
    #s-content.s-font-heading-source-sans-pro .s-font-heading:lang(zh) {
 
      font-family: "source sans pro", helvetica, 'PingFang SC',"Microsoft YaHei","微软雅黑",STXihei,"华文细黑", sans-serif
 
    }
 
    #s-content.s-font-heading-source-sans-pro .s-font-heading:lang(zh-tw) {
 
      font-family: "source sans pro", helvetica, 'Lisong Pro', 'PMingLiU', 'PingFang TC','Microsoft JhengHei',"微軟正黑體",STXihei, sans-serif;
 
    }
 
  </style></div><div name="topBar" type="NavTheme" id="f_a7eefaef-c78a-4fe1-925d-f515062961c4" class="css-NavbarSection-192rqjg0 s-nav navigator css-3109jj"><div class="s-navbar-desktop s-navbar-desktop-normal s-bg-dark-text"><div class="s-nav-inner"><div class="s-logo"><div class="css-LogoImageContainerDiv-192rqjg1 css-enscdp"><div class="s-logo-image-normal"><div class="s-logo-image image1"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1518694/RDFZ_logo_v1qehu.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div></div><div class="s-logo-image-alternative"><div class="s-logo-image image1"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1518694/RDFZ_logo_v1qehu.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div><div class="s-logo-image image2"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1134318/logo_a7pca6.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div></div></div></div><div class="s-nav-items-and-links"><ul class="s-uncollapsed-nav "><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#home">HOME</a></div></li><li style="display:none" class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#boards">BOARDS</a></div></li><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#safety">Safety</a></div></li><li class="s-nav-li s-nav-ellipsis hidden s-navbar-dropdown _down"><div class="s-nav-link-container"><a><span class="s-font-body">… <!-- --> <i style="display:inline" class="fa fa-angle-down"></i></span></a></div><ul class="s-collapsed-nav"><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#home">HOME</a></div></li><li style="display:none" class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#boards">BOARDS</a></div></li><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#safety">Safety</a></div></li></ul></li></ul></div><ul class="s-nav-icons"><li class="s-nav-li"></li></ul></div></div><div class="s-navbar-desktop s-navbar-desktop-fixed strikingly-fixed s-bg-dark-text _alternativeNavBackgroundColorIsDifferentFromNormal"><div class="s-nav-inner"><div class="s-logo"><div class="css-LogoImageContainerDiv-192rqjg1 css-enscdp"><div class="s-logo-image-normal"><div class="s-logo-image image1"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1518694/RDFZ_logo_v1qehu.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div></div><div class="s-logo-image-alternative"><div class="s-logo-image image1"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1518694/RDFZ_logo_v1qehu.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div><div class="s-logo-image image2"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1134318/logo_a7pca6.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div></div></div></div><div class="s-nav-items-and-links"><ul class="s-uncollapsed-nav "><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#home">HOME</a></div></li><li style="display:none" class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#boards">BOARDS</a></div></li><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#safety">Safety</a></div></li><li class="s-nav-li s-nav-ellipsis hidden s-navbar-dropdown _down"><div class="s-nav-link-container"><a><span class="s-font-body">… <!-- --> <i style="display:inline" class="fa fa-angle-down"></i></span></a></div><ul class="s-collapsed-nav"><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#home">HOME</a></div></li><li style="display:none" class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#boards">BOARDS</a></div></li><li class="s-nav-li"><div class="s-nav-link-container"><a class="s-font-body" href="#safety">Safety</a></div></li></ul></li></ul></div><ul class="s-nav-icons"><li class="s-nav-li"></li></ul></div></div><div class="s-navbar-mobile-header"><div class="s-logo-image image1"><div class="s-component s-image"><div><div class="s-component-content"><div><img src="//res.cloudinary.com/hrscywv4p/image/upload/c_limit,fl_lossy,h_300,w_300,f_auto,q_auto/v1/1518694/RDFZ_logo_v1qehu.png" alt="" title="" class="crop-default" data-description=""/></div></div></div></div></div></div></div><div class="css-S5PageWrapperDiv-evtmtg0 css-1c86csr"><div class="s-navbar-container"><div class="navbar-drawer-bar"><div class="drawer-container"><a class="navbar" href="javascript:void(0);"><div class="navbar-icons"><span></span><span></span><span></span></div></a>
 
      <!-- -->
 
      <div class="mobile-cart-icon" id="s-mobile-shopping-cart-new"></div><div class="navbar-drawer-title-container"></div></div></div><div id="navbar-drawer-mask"></div><div class="navbar-drawer strikingly-drawer bottom-border"><ul id="nav-drawer-list"><span><li><a class="navbar-drawer-item s-font-body" href="#1">HOME</a></li></span><span><li style="display:none"><a class="navbar-drawer-item s-font-body" href="#2">BOARDS</a></li></span><span><li><a class="navbar-drawer-item s-font-body" href="#3">Safety</a></li></span><li id="mobileLoginDropdown"></li><li class="logo"><a class="logo" href="https://www.strikingly.com/?ref=logo&amp;permalink=site-1518694-4764-3608&amp;m=1" target="_blank"><div class="msg">Powered By</div><img src="https://static-assets.strikinglycdn.com/images/logo-small-2.png" style="height:28px" alt="Strikingly"/></a></li></ul></div><div class="mobile-actions"><ul id="mobile-action-list"></ul></div></div><div class="page-wrapper"><div id="prerendered-inline-theme-html"></div><ul style="display:block" class="slides s-page-1"><li class="slide s-section-1 s-first-visible-section css-1nxcohz" id="section-f_4b871ddc-bcab-465e-a9ed-89bc2f5426a6"><div class="waypoint"></div><a class="section-anchor"></a><div>
 
  <div class="lazyload  s-no-bg s-bg-white s-new-hero-section s-section s-hero-section" data-react-style="{}">
 
  <div class="container s-rva s-layout-buttonLeft"><div class="s-rva-media eight columns  empty-media"><div class="s-component s-media "><div></div></div></div><div class="s-rva-text eight columns "><div class="half-offset-right"><div class="s-title-group  "><div class="s-title sb-title-left-align"><div class="s-component s-text"><h1 class="s-component-content s-font-title"><p style="font-size: 130%; text-align: left;"><span class="s-text-color-black"><strong>Xscape</strong></span></p></h1></div></div><div class="s-subtitle"><div class="s-component s-text"><h4 class="s-component-content s-font-heading"><p style="text-align: left; font-size: 130%;"><span class="s-text-color-black">Rational designed biosafety device for fermentation and therapy</span></p></h4></div></div></div></div></div></div>
 
</div>
 
</div></li><li class="slide s-section-2 css-10c7nxj" id="section-f_4694f3bf-c87c-44d4-9119-4c6cd50cde38"><div class="waypoint"></div><a class="section-anchor"></a><div>
 
  <div class="lazyload  s-bg-white s-new-media-section s-section s-columns-section  " data-react-style="{}"><div class="container"><div class="columns sixteen"><div class="s-title-group  "><div class="s-subtitle"><div class="s-component s-text"><h4 class="s-component-content s-font-heading"><p style="font-size: 160%;" class="s-text-color-black"><b>Devices</b></p></h4></div></div></div></div>
 
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Measurement

Expression Burden

In synthetic biology, a common issue in integrating exogenous sequences into host cells is that heterologous gene expression can cause significant metabolic burden. Typically, this gene expression burden is implied by a declining growth rate of bacteria, which is unfavorable for fermentation industry because it lowers yields and profits. According to Ceroni et al. (2015), a fluorescence-based method allows to measure gene expression capacity of cells, by inserting a “capacity monitor”.

Capacity Monitor

The “Capacity Monitor” consists of a superfold GFP with a strong constitutive promoter, which is constantly expressed in the bacterial genome. Thus, any alternation in fluorescence intensity can reflect in overall resource availability in the cell. Particularly, with expression of synthesized genes, the expression of GFP will decrease, thus reporting a reallocation of cellular resources.

VioABDE

After applying capacity monitor, we compared the fluorescence of E. coli with an empty vector and E. coli with VioABDE inserted and expressed. Both strains were incubated for 24 hours with fluorescence and OD constantly monitored. However, the OD of bacteria with VioABDE increased faster than the bacteria without Vio expression, which contradicts our initial expectation.

We co transformed the TlpA39-VioABDE into the capcatiy monitor component cell which was made by beyotime component cell kit.

Then the Capacity Monitor and Capacity Monitor-Vio were incubicated in microshaker in 39 degree Celsius, then they were added to the 96 well plates. "

Alternative Device

Since we target to modify bacteria used in fermentation industry, which primarily are used for material synthesis and production, our sequence inserts should occupy as limited resources in bacteria as possible to avoid significant expression burden. In order to reduce this expression stress, we designed another device for fermentation which used a LuxR repressive promoter (Peking-S, 2011) and the “Cold Box” in 5’UTR region of CspA (Ionis Paris, 2017). With only one transcriptional regulator, less energy will be consumed, which is beneficial for fermentation industry.

References

Ceroni, F., Algar, R., Stan, G. B., & Ellis, T. (2015). Quantifying cellular capacity identifies gene expression designs with reduced burden. Nature methods, 12(5), 415.

Ionis Paris. (2017). Biological thermo-responsive solution, that reduces the plant’s thermal stresses. Available from: https://2017.igem.org/Team:IONIS-PARIS.

Peking-S iGEM. (2011). A Chemical Wire Toolbox for Synthetic Microbial Consortia. Avaible from: https://2011.igem.org/Team:Peking_S.

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