Difference between revisions of "Team:Exeter/Composite Part"

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<div class="column full_size judges-will-not-evaluate">
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<h3>★  ALERT! </h3>
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<p>This page is used by the judges to evaluate your team for the <a href="https://2017.igem.org/Judging/Medals">medal criterion</a> or <a href="https://2017.igem.org/Judging/Awards"> award listed above</a>. </p>
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<p> Delete this box in order to be evaluated for this medal criterion and/or award. See more information at <a href="https://2017.igem.org/Judging/Pages_for_Awards"> Instructions for Pages for awards</a>.</p>
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<!-- Row & container for main content of the page -->
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<div class="container-fluid">
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      <a href="https://2017.igem.org/Team:Exeter"><img id="logo-top-left" width="100%;" src="https://static.igem.org/mediawiki/2017/2/29/T--Exeter--startpage_logo.png"></a>
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      <nav id="navbar-page" class="navbar"> <!-- classes removed from template: navbar-light bg-light-->
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        <!-- <a class="navbar-brand" href="#navbar-top">Back to top</a> -->
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        <nav class="nav nav-pills flex-column">
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          <a class="nav-link" href="#h1">FimH sfGFP</a>
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          <a class="nav-link" href="#h2">FimH Fusion Proteins</a>
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          <a class="nav-link" href="#h3">Fim Operon</a>
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          <a class="nav-link" href="#pageHeader">back to top</a>
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    <div class="col-10" id="pageContent">
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        <img class="w-25 mx-auto d-block" src="https://static.igem.org/mediawiki/2017/a/a4/T--Exeter--Composite_part_logo.png">
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        <h1 id="pageHeader">Composite Parts</h1>
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          <p>We have decided to use five different promoters to drive expression of our pili proteins. The T7 promoter (BBa_K1614000) was chosen as it is a strong, well characterised promoter but as this could only be used in BL21(DE3) it was only used for initial expression studies. Three inducible promoters were chosen to allow control over expression in non pili-producing strains of <i>E. coli</i>: the rhamnose inducible promoter P_Rha (BBa_K902065); the arabinose inducible promoter P_ara (BBa_I13453) and the IPTG inducible T5 promoter (pQE30-Qiagen). Finally the constitutive promoter P_J23100 (BBa_J23100) was chosen.</p>
  
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<p>We chose the well characterised strong RBS BBa_B0034 and the double terminator BBa_B0015.</p>
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<h1>Composite Parts</h1>
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<p>All parts were synthesised as gBlocks by IDT and composite parts were built using a one-pot cloning method <a href="https://2017.igem.org/Team:Exeter/Experiments"> protocol</a> modified from Weber et al 2011. All FimH constructs were inserted into pSB1A3 and the fim operon constructs were inserted into pX1’6’0’0 (lab built plasmid). Despite multiple efforts not all of the planned constructs could be built, but the ones that were successful, listed below, were submitted to iGEM in pSB1C3.</p>
  
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<p>
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<h2 id="h1">FimH sfGFP</h2>
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A composite part is a functional unit of DNA consisting of two or more basic parts assembled together. <a href="http://parts.igem.org/wiki/index.php/Part:BBa_I13507">BBa_I13507</a> is an example of a composite part, consisting of an RBS, a protein coding region for a red fluorescent protein, and a terminator.
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</p>
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<p>New composite BioBrick devices can be made by combining existing BioBrick Parts (like Inverters, Amplifiers, Smell Generators, Protein Balloon Generators, Senders, Receivers, Actuators, and so on).</p>
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<table style="width:90%; margin:0 auto; border-collapse: collapse;  border: 2px solid black" border="1" >
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  <tr>
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<th>Name</th>
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<th>Description</th>
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<th>Base Pairs</th>
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  </tr>
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  <tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324007">BBa_K2324007</a></td>
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<td>P_Rha_FimH_1_sfGFP</td>
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<td>2020</td>
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  </tr>
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  <tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324006">BBa_K2324006</a></td>
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<td>P_Rha_FimH_225_sfGFP</td>
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<td>2020</td>
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  </tr>
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<tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324008">BBa_K2324008</td>
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<td>P_Rha_FimH_258_sfGFP</td>
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<td>2020</td>
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  </tr>
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                                <tr>
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                                      <td><a href="http://parts.igem.org/Part:BBa_K2324011">BBa_K2324011</td>
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                                      <td>P_T7_FimH_225sfGFP</td>
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                                      <td>1798</td>
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</table>
  
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        <h2 id="h2">FimH Fusion Proteins</h2>
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<h3>Best Composite Part Special Prize</h3>
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<table style="width:90%; margin:0 auto; border-collapse: collapse;  border: 2px solid black" border="1" >
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  <tr>
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<th>Name</th>
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<th>Description</th>
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<th>Base Pairs</th>
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  </tr>
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  <tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324013">BBa_K2324013</a></td>
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<td>P_Rha_FimH_1_His</td>
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<td>1102</td>
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  </tr>
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  <tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324009">BBa_K2324009</a></td>
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<td>P_Rha_FimH_1_SynMT</td>
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<td>1474</td>
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  </tr>
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<tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324010">BBa_K2324010</td>
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<td>P_Rha_FimH_1_MouseMT</td>
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<td>1489</td>
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  </tr>
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</table>
  
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<p>New BioBrick devices can be made by combining existing BioBrick Parts. For example, Inverters, Amplifiers, Smell Generators, Protein Balloon Generators, Senders, Receivers, Actuators, and so on. To be eligible for this award, this part must adhere to <a href="http://parts.igem.org/DNA_Submission">Registry sample submission guidelines</a> and have been sent to the Registry of Standard Biological Parts. If you have a part you wish to nominate your team for this <a href="https://2017.igem.org/Judging/Awards">special prize</a>, make sure you add your part number to your <a href="https://2017.igem.org/Judging/Judging_Form">judging form</a> and delete the box at the top of this page.
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        <h2 id="h3">Fim Operon</h2>
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<table style="width:90%; margin:0 auto; border-collapse: collapse;  border: 2px solid black" border="1" >
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  <tr>
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<th>Name</th>
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<th>Description</th>
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<th>Base Pairs</th>
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  <tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324012">BBa_K2324012</a></td>
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<td>P_J23100_Fim_Operon</td>
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<td>5944</td>
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  </tr>
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  <tr>
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<td><a href="http://parts.igem.org/Part:BBa_K2324015">BBa_K2324015</a></td>
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<td>P_Ara_Fim_Operon</td>
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<td>5874</td>
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  </tr>
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</table>
  
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<h2>Reference</h2>
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<b>Please note:</b> Judges will only look at the first part number you list, so please only enter ONE (1) part number in the judging form for this prize. </p>
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<p id="referenceList">Weber, E., Engler, C., Gruetzner, R., Werner, S., and Marrillonnet, S. (2011) A Modular Cloning System for Standardized Assembly of Multigene Constructs. PLOS One 6, e16765</p>
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<div class="highlight">
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<h4>Note</h4>
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<p>This page should list all the composite parts your team has made during your project. You must add all characterization information for your parts on the Registry. You should not put characterization information on this page. Remember judges will only look at the first part in the list for the Best Composite Part award, so put your best part first!</p>
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{{Exeter/footer}}

Latest revision as of 21:41, 1 November 2017

Composite Parts

We have decided to use five different promoters to drive expression of our pili proteins. The T7 promoter (BBa_K1614000) was chosen as it is a strong, well characterised promoter but as this could only be used in BL21(DE3) it was only used for initial expression studies. Three inducible promoters were chosen to allow control over expression in non pili-producing strains of E. coli: the rhamnose inducible promoter P_Rha (BBa_K902065); the arabinose inducible promoter P_ara (BBa_I13453) and the IPTG inducible T5 promoter (pQE30-Qiagen). Finally the constitutive promoter P_J23100 (BBa_J23100) was chosen.

We chose the well characterised strong RBS BBa_B0034 and the double terminator BBa_B0015.

All parts were synthesised as gBlocks by IDT and composite parts were built using a one-pot cloning method protocol modified from Weber et al 2011. All FimH constructs were inserted into pSB1A3 and the fim operon constructs were inserted into pX1’6’0’0 (lab built plasmid). Despite multiple efforts not all of the planned constructs could be built, but the ones that were successful, listed below, were submitted to iGEM in pSB1C3.

FimH sfGFP

Name Description Base Pairs
BBa_K2324007 P_Rha_FimH_1_sfGFP 2020
BBa_K2324006 P_Rha_FimH_225_sfGFP 2020
BBa_K2324008 P_Rha_FimH_258_sfGFP 2020
BBa_K2324011 P_T7_FimH_225sfGFP 1798

FimH Fusion Proteins

Name Description Base Pairs
BBa_K2324013 P_Rha_FimH_1_His 1102
BBa_K2324009 P_Rha_FimH_1_SynMT 1474
BBa_K2324010 P_Rha_FimH_1_MouseMT 1489

Fim Operon

Name Description Base Pairs
BBa_K2324012 P_J23100_Fim_Operon 5944
BBa_K2324015 P_Ara_Fim_Operon 5874

Reference

Weber, E., Engler, C., Gruetzner, R., Werner, S., and Marrillonnet, S. (2011) A Modular Cloning System for Standardized Assembly of Multigene Constructs. PLOS One 6, e16765