Difference between revisions of "Team:H14Z1 Hangzhou/Design"

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<h1>Design</h1>
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Design is the first step in the design-build-test cycle in engineering and synthetic biology. Use this page to describe the process that you used in the design of your parts. You should clearly explain the engineering principles used to design your project.
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This page is different to the "Applied Design Award" page. Please see the <a href="https://2018.igem.org/Team:H14Z1_Hangzhou/Applied_Design">Applied Design</a> page for more information on how to compete for that award.
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<h3>What should this page contain?</h3>
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    <div id="page_content_container">
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<ul>
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        <div id="menu" attribute="Description"></div>
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<li>Explanation of the engineering principles your team used in your design</li>
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<li>Discussion of the design iterations your team went through</li>
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<li>Experimental plan to test your designs</li>
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</ul>
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        <div class="content">
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            <img src="https://static.igem.org/mediawiki/2018/4/49/T--H14Z1_Hangzhou--head_overview.png" alt="" class="head_div_img" />
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            <div class="content_box">
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<div class="highlight decoration_A_full">
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                <h1 class="content_title">Design</h1>
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<h3>Inspiration</h3>
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                <div class="content_conts">
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<ul>
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                    <!------------------------Liver function and protection------------------------------ -->
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<li><a href="https://2016.igem.org/Team:MIT/Experiments/Promoters">2016 MIT</a></li>
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                    <h3 class="content_subtitle">Liver function and protection</h3>
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<li><a href="https://2016.igem.org/Team:BostonU/Proof">2016 BostonU</a></li>
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                    <p class="content_context"  style="text-indent:2em; text-align:justify">
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<li><a href="https://2016.igem.org/Team:NCTU_Formosa/Design">2016 NCTU Formosa</a></li>
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                        <a href="http://www.discoverymedicine.com/category/species-and-cell-types/bacterium/lactococcus-lactis/">Lactococcus lactis</a>  
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                        is one of the most common organisms used in bacterial drug delivery system.
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                        <a href="http://www.discoverymedicine.com/category/species-and-cell-types/bacterium/lactococcus-lactis/">Lactococcus lactis</a>  
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</div>
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                        is a food-grade bacterium that is widely used in the dairy industry. It is a
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                        Gram-positive bacterium and therefore lacks endotoxic lipopolysaccharides (LPS) which are
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                        associated with commonly used
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                        <a href="http://www.discoverymedicine.com/category/species-and-cell-types/bacterium/gram-negative-bacteria/">Gram-negative bacteria</a>
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                        . It is also non-invasive and non-commensal
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                        and thus has less potential to trigger immunotolerance or side effects upon prolonged use. The
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                        bacterium has an established safety profile through its long use in fermented dairy products
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                        and is considered as a GRAS (generally recognized as safe) microorganism. During the last two
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                        decades, significant advances have been made in the field of lactococcal genetics and protein
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                        expression systems. The most commonly used system is the nisin-controlled gene expression
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                        (NICE) system, containing the nisin promoter. Consequently, we have seen the emergence of new
 +
                        areas for the application of engineered L. lactis for protein expression,
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                        <a href="http://www.discoverymedicine.com/category/research-technology/nucleic-acid/gene-delivery/">gene delivery</a>,
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                        vaccine delivery, and therapeutic
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                        <a href="http://www.discoverymedicine.com/category/research-technology/drug-delivery-research-technology/">drug delivery.</a>
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                    </p>
  
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                    <p class="content_context" style="text-indent:2em; text-align:justify">
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                        In the present project, since the oral table administration of GSH and SAM have some
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                        disadvantages, such as low stability and short life span, here we tried to develop a novel
 +
                        in-vivo strategy of produce and deliver them simultaneously by using NICE system. In the
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                        experiment, two-functional GSH synthetase gene (gshF) and SAM synthetase gene (metK) were in
 +
                        tandem inserted into the expression vector (pNZ8148), and the resulted plasmid (pNZ8148-SG) was
 +
                        employed to construct the target vector pNZ8148-SGC by introducing adhesion factor gene (cwaA).
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                        This target vector was transformed to get recombinant Lactococcus lacti, which was employed to
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                        produce our “smart yogurt”.
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                    </p>
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                    <h3 class="content_subtitle">
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                        The project is outlined with two stages：
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                    </h3>
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                    <!-- ------------------------Stage A---------------------------- -->
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                    <p class="mt8 content_context">
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                        <span style="width:120px; font: bold; font-size: 25px">Stage A</span><span>Construction of Lactococcus lactis NZ9000/pNZ-SGC</span>
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                    </p>
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                    <p><img style="width: 50%; margin-top: 1em" src="https://static.igem.org/mediawiki/2018/b/be/T--H14Z1_Hangzhou--description_projectOverview_fig1.png"></p>
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                    <p><img style="width: 50%; margin-top: 1em" src="https://static.igem.org/mediawiki/2018/c/cd/T--H14Z1_Hangzhou--description_projectOverview_fig12.png"></p>
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                    <p><img style="width: 50%; margin-top: 1em" src="https://static.igem.org/mediawiki/2018/8/87/T--H14Z1_Hangzhou--description_projectOverview_fig3.png"></p>
  
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                    <!-- ------------------------Stage B---------------------------- -->
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                    <p class="mt8 content_context">
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                        <span style="width:120px; font: bold; font-size: 25px">Stage B</span><span>Preparation of smart yogurt with our constructed Lactococcus lactis</span>
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                    </p>
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                    <p><img style="width: 50%; margin-top: 1em" src="https://static.igem.org/mediawiki/2018/6/66/T--H14Z1_Hangzhou--description_projectOverview_fig4.png"></p>
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Revision as of 23:05, 13 October 2018

<!DOCTYPE html>

Design

Liver function and protection

Lactococcus lactis is one of the most common organisms used in bacterial drug delivery system. Lactococcus lactis is a food-grade bacterium that is widely used in the dairy industry. It is a Gram-positive bacterium and therefore lacks endotoxic lipopolysaccharides (LPS) which are associated with commonly used Gram-negative bacteria . It is also non-invasive and non-commensal and thus has less potential to trigger immunotolerance or side effects upon prolonged use. The bacterium has an established safety profile through its long use in fermented dairy products and is considered as a GRAS (generally recognized as safe) microorganism. During the last two decades, significant advances have been made in the field of lactococcal genetics and protein expression systems. The most commonly used system is the nisin-controlled gene expression (NICE) system, containing the nisin promoter. Consequently, we have seen the emergence of new areas for the application of engineered L. lactis for protein expression, gene delivery, vaccine delivery, and therapeutic drug delivery.

In the present project, since the oral table administration of GSH and SAM have some disadvantages, such as low stability and short life span, here we tried to develop a novel in-vivo strategy of produce and deliver them simultaneously by using NICE system. In the experiment, two-functional GSH synthetase gene (gshF) and SAM synthetase gene (metK) were in tandem inserted into the expression vector (pNZ8148), and the resulted plasmid (pNZ8148-SG) was employed to construct the target vector pNZ8148-SGC by introducing adhesion factor gene (cwaA). This target vector was transformed to get recombinant Lactococcus lacti, which was employed to produce our “smart yogurt”.

The project is outlined with two stages：

Stage AConstruction of Lactococcus lactis NZ9000/pNZ-SGC

Stage BPreparation of smart yogurt with our constructed Lactococcus lactis