Difference between revisions of "Team:Amazonas Brazil/Project"

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       <ul class="Links_bar" style="background: linear-gradient(to left, #263D9E, #3F2759);">
 
         <li><a class="Link-pg" href="https://2017.igem.org/Team:Amazonas_Brazil"><img src="https://static.igem.org/mediawiki/2017/d/d4/T--Amazonas_Brazil--LogoBranco.png"  width="104.04";height="90"; class="Menu_image"></a></li>
 
         <li><a class="Link-pg" href="https://2017.igem.org/Team:Amazonas_Brazil"><img src="https://static.igem.org/mediawiki/2017/d/d4/T--Amazonas_Brazil--LogoBranco.png"  width="104.04";height="90"; class="Menu_image"></a></li>
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         <li><a class="bar_text" href="https://2017.igem.org/Team:Amazonas_Brazil/Demonstrate">Demonstrate</a></li>
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         <li><a class="bar_text" href="https://2017.igem.org/Team:Amazonas_Brazil/Project">Description</a></li>
 
         <li><a class="bar_text" href="https://2017.igem.org/Team:Amazonas_Brazil/Model">Modelling</a></li>
 
         <li><a class="bar_text" href="https://2017.igem.org/Team:Amazonas_Brazil/Model">Modelling</a></li>
 
         <li><a class="bar_text" href="https://2017.igem.org/Team:Amazonas_Brazil/InterLab">Interlab</a></li>
 
         <li><a class="bar_text" href="https://2017.igem.org/Team:Amazonas_Brazil/InterLab">Interlab</a></li>
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<div class="hero-full-container background-image-container white-text-container" style="background-image: url(https://static.igem.org/mediawiki/2017/1/11/T--Amazonas_Brazil--PROJECT.jpg); " >
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  <div class="hero-full-container background-image-container white-text-container" style="background: linear-gradient(to left, #263D9E, #3F2759);">
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  <div class="container">
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    <div class="container">
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        <div class="col-xs-12">
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          <div class="text-content  text-center">
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            <div class="text-content  text-center">
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            <h2 class="Page_Title" align="middle">PROJECT</h2>
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              <h2 class="Page_Title" align="middle">DEMONSTRATE</h2>
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          <h2 class="Page_Subtitle" align="middle">pCRISPeasy</h2>
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             <h1 class="First"style="text-align: center;height: 2.3em;">Navigate into CRISPeasy roadmap</h1>
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             <h1 class="First"style="text-align: center;">Demonstrate</h1>
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              <div >
  
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                <img style="display: block; margin-left: auto; margin-right: auto;width: 26%;height: 20%"
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                src="https://static.igem.org/mediawiki/2017/0/02/Amazonas_Brazil_2017Demonstrate1.png">
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      <!--      </div> -->  
  
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              <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto"
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                <div style='position:absolute;top:100%; left:6%;'>                 
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                          src="https://static.igem.org/mediawiki/2017/a/ab/Fluxo.png"/>
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                <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto; width:45%; height:45%;"
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                src="https://static.igem.org/mediawiki/2017/8/87/Amazonas_Brazil_2017OPERATION1.png"/>                 
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                </div>
  
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                <div style='position:absolute;top:47%; left:45%;'> <!-- div para imagens que se sobrepoem -->
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                <!--
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                  <a href="https://2017.igem.org/Team:Amazonas_Brazil/Human_Practices">
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                <div style='position:absolute;top:53%; left:4%;'>                
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                      <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto; width:150px; height:50px;"
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                <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto; width:45%; height:45%;"
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                                              src="https://static.igem.org/mediawiki/2017/1/1a/1_HumanPratices.png"/>
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                src="https://static.igem.org/mediawiki/2017/0/0d/Amazonas_Brazil_2017CROUT1.png"/>                
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                  </a>
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                      <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto"
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                src="https://static.igem.org/mediawiki/2017/1/12/Amazonas_Brazil_2017CRIN1.png"/>                
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                                              src="https://static.igem.org/mediawiki/2017/3/37/2_Specifications.png"/>
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                <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto; width:45%; height:45%;"
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                  <a href="#Design">
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                src="https://static.igem.org/mediawiki/2017/0/02/Amazonas_Brazil_2017Demonstrate1.png"/>                   
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                      <img alt="Erro" style="display: block; margin-left: auto; margin-right: auto"
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                </div>
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                  <a href="https://2017.igem.org/Team:Amazonas_Brazil/Model">
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                                              src="https://static.igem.org/mediawiki/2017/7/79/4_Modelling.png"/>
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                  <a href="https://2017.igem.org/Team:Amazonas_Brazil/Demonstrate#Assembly">
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                                              src="https://static.igem.org/mediawiki/2017/4/4c/5_Assembly.png"/>
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                <div style='position:absolute;top:66%; left:25%;'> <!-- div para imagens que se sobrepoem -->
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                  <a href="https://2017.igem.org/Team:Amazonas_Brazil/Demonstrate#Testing">
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                                              src="https://static.igem.org/mediawiki/2017/a/a4/6_Testing.png"/>
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                  <a href="https://2017.igem.org/Team:Amazonas_Brazil/Applied_Design">
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                                              src="https://static.igem.org/mediawiki/2017/3/38/7_Implementation.png"/>
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               <h1 style=" margin: 0;padding-top: 0.1em;"id="Specifications"></h1>
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              <h2 style="margin-top: 1.4em;">Specifications</h2>
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              <h3>Our proposal is to provide a bacterial genome editing approach based on BioBrick parts assembly easy to engineer as A, B, C… CRISPR</h3>
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              <p class="p">
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                The scientific community had achieved great results using CRISPR-Cas9 mediated genome editing techniques. While these methods are moving forward, some basic methodological aspects related to this approach could be further improved to expand this scientific revolution.
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In prokaryotes, the genome editing based on CRISPR/Cas9 machinery require quite laborious days of work, the system is weakly founded on bioengineering principles, demands lots of steps, each one carrying a part of the CRISPR/Cas9 apparatus. In addition, off-target events happen frequently, the machinery post-effect in the cell isn’t entirely known yet, among others. To legitimate this information, we integrated our project design with other iGEM's teams who had worked with CRISPR/Cas9 in previous years. You can read more about what we found here (LINK PRA INTEGRADA)
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The decoupling of all these shortcomings paved the way to the project's delineation and we elected to focus on a challenge: the lack of standardization, which is a real-world problem - included but not limited to SynBio. Then, to optimize the previous methods, we designed a trustworthy, well functioning standardized framework based on BioBrick parts called: the CRISPeasy toolbox.
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                </p>
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              <div>
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            </div>
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                <h1 style=" margin: 0;padding-top: 0.1em;"id="Assembly"></h1>
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            <div>
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                <h2 style="margin-top: 1.4em;">Toolbox modules assembly</h2>
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            <h1 style=" margin: 0;padding-top: 0.1em;" id="Design" ></h1>
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                <h4 style="padding: 0;">How we constructed our circuits</h4>
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            <h2 style="margin-top: 1.4em;">Design</h2>
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                <h3 style="padding: 0;margin-top: 0.5em;margin-bottom: 0.5em;">core breakthroughs</h3>
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            <h3>Check all the parts involved and our approaches with closer details!</h3>
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                 <li>Construction of standard interchangeable parts for genome engineering based on CRISPR-Cas9 machinery;</li>
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              <p class="p">
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                 <li>Engineering of an optimized sgRNA (both crRNA design AND tracrRNA structure) to amplify editing efficiency;</li>
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                 Cas9 (BBa_K2457001): Standardized Cas9 device, regulated by the AraC_pBAD regulatory module.
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              </div>
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                 In the single guide RNA presence, it forms a catalytically active ribonucleoprotein (RNP) complex
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              <div>
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                which cleaves specifics target sequences from DNA, leading to double-strand breaks. Through cell repair
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               <h1 style=" margin: 0;padding-top: 0.1em;"id="Testing"></h1>
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                pathways, it will lays the road to genomic editing in living cells.
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                <h2 style="margin-top: 1.4em;">CRISPeasy-out method (crOUT)</h2>
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              </p>
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                <h3 style="padding: 0;margin-top: 0.5em;margin-bottom: 0.5em;">core breakthroughs:</h3>
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            </div>
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                <li>Characterization of the Cas9 device + optimized sgRNA (BBa_K2457006) in E. coli K12 MG1655 cells by tracking the impact on the cellular system (i.e., growth ratio);</li>
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            <div>
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                <li>Setting up a reliable data sheet of Cas9 activation machinery in a range of conditions to further expand its uses by SynBio and iGEM community;</li>
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               <img src="https://static.igem.org/mediawiki/2017/5/5f/T--Amazonas_Brazil--infografico_project.png" style="background-size: cover; background-repeat: no-repeat;width:80%; margin-left: 10%;">
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              </div>
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            </div>
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              <div style="background-color:#323232"> <!-- Div para imagens-->
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                <div>
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                  <p>
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                    Teste
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                  </p>
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Revision as of 02:45, 1 November 2017

Wiki_iGEM_Amazonas

DEMONSTRATE

Demonstrate

Erro

Toolbox modules assembly

How we constructed our circuits

core breakthroughs

  • Construction of standard interchangeable parts for genome engineering based on CRISPR-Cas9 machinery;
  • Engineering of an optimized sgRNA (both crRNA design AND tracrRNA structure) to amplify editing efficiency;

    • CRISPeasy-out method (crOUT)

    core breakthroughs:

  • Characterization of the Cas9 device + optimized sgRNA (BBa_K2457006) in E. coli K12 MG1655 cells by tracking the impact on the cellular system (i.e., growth ratio);
  • Setting up a reliable data sheet of Cas9 activation machinery in a range of conditions to further expand its uses by SynBio and iGEM community;

    • Teste