Difference between revisions of "Team:Edinburgh/Parts"

Line 1: Line 1:
−
{{Edigem15_wikireset}}
+
{{Edinburgh_Basic}}
 
<html>
 
<html>
  
−
<h2> Part Documentation</h2>
 
  
−
<p>Each team will make new parts during iGEM and will submit them to the Registry of Standard Biological Parts. The iGEM software provides an easy way to present the parts your team has created. The <code>&lt;groupparts&gt;</code> tag (see below) will generate a table with all of the parts that your team adds to your team sandbox.</p>
+
    <head>
−
<p>Remember that the goal of proper part documentation is to describe and define a part, so that it can be used without needing to refer to the primary literature. Registry users in future years should be able to read your documentation and be able to use the part successfully. Also, you should provide proper references to acknowledge previous authors and to provide for users who wish to know more.</p>
+
<link rel="stylesheet" href="https://maxcdn.bootstrapcdn.com/bootstrap/3.3.5/css/bootstrap.min.css">
 +
<script src="https://maxcdn.bootstrapcdn.com/bootstrap/3.3.5/js/bootstrap.min.js"></script>
 +
    </head>
 +
<!-- menu -->
 +
<div id="custom-bootstrap-menu" class="navbar navbar-default navbar-fixed-top" role="navigation">
 +
    <div class="container-fluid">
 +
        <div class="navbar-header">
 +
            <button type="button" class="navbar-toggle" data-toggle="collapse" data-target=".navbar-menubuilder"><span class="sr-only">Toggle navigation</span><span class="icon-bar"></span><span class="icon-bar"></span><span class="icon-bar"></span>
 +
            </button>
 +
        </div>
 +
        <div class="collapse navbar-collapse navbar-menubuilder">
 +
            <ul class="nav navbar-nav navbar-right">
 +
              <li class="active">
 +
                  <a href="https://2015.igem.org/Team:Edinburgh">Home</a></li>
 +
                  <li class="dropdown">
 +
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">About<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Team">Team</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Attributions">Attributions</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Collaborations">Collaboration</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Sponsors">Sponsors</a></li>
  
−
 
+
                    </ul>
−
<div class="highlightBox">
+
                  </li>
−
<h4>Note</h4>
+
                  <li class="dropdown">
−
<p>Note that parts must be documented on the <a href="http://parts.igem.org/Main_Page"> Registry</a>. This page serves to <i>showcase</i> the parts you have made. Future teams and other users and are much more likely to find parts by looking in the Registry than by looking at your team wiki.</p>
+
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">Project<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Description">Overview</a></li>
 +
                    <!-- <li><a href="https://2015.igem.org/Team:Edinburgh/Experiments">Experiments</a> </li> -->
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/HeroinBiosensor">Heroin Biosensor</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/DNPBiosensor">DNP Biosensor</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/PMABiosensor">PMA Biosensor</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/CBD">Making it Stick</a></li>             
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Results">Results</a></li>
 +
                    </ul>
 +
                  </li>
 +
                  <li class="dropdown">
 +
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">Parts<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Parts">Team Parts</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Basic_Part">Basic Parts</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Composite_Part">Composite Parts</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Part_Collection">Part Collection</a> </li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Improved_Part">Improved Parts</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Characterisation_Part">Improved Characterisation</a></li>
 +
                    </ul>
 +
                  </li>
 +
                  <li class="dropdown">
 +
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">Dry Lab<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Modeling">Modeling</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Design">Design</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Software">Software</a></li>
 +
                    </ul>
 +
                  </li>
 +
                  <li class="dropdown">
 +
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">Human Practices<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Practices">Our Story</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Safety">Safety</a> </li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Legality">Legality</a> </li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Outreach">Outreach</a> </li>
 +
                    </ul>
 +
                  </li>
 +
                  <li class="dropdown">
 +
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">InterLab<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/InterLab">InterLab</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/InterLabBook">LabBook</a> </li>
 +
                    </ul>
 +
                  </li>
 +
                  <li class="dropdown">
 +
                    <a href="#" class="dropdown-toggle" data-toggle="dropdown" role="button" aria-expanded="false">Notebook<span class="caret"></span></a>
 +
                    <ul class="dropdown-menu" role="menu">
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Project/Protocols">Protocols</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Notebook/HeroinPurity">Heroin Purity</a></li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Notebook/PMADetection">PMA Detection</a> </li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Notebook/DNPDetection">DNP Detection</a> </li>
 +
                      <li><a href="https://2015.igem.org/Team:Edinburgh/Notebook/FluidDynamics">Fluid Dynamics</a> </li>
 +
                    </ul>
 +
                  </li>
 +
                  <li><a href="https://2015.igem.org/Team:Edinburgh/MedalCriteria">Medal Criteria</a></li> 
 +
            </ul>
 +
        </div>
 +
    </div>
 
</div>
 
</div>
  
  
 +
    <!-- End of menu  -->
  
−
<h4>Adding parts to the registry</h4>
+
<body>
−
<p>You can add parts to the Registry at our <a href="http://parts.igem.org/Add_a_Part_to_the_Registry">Add a Part to the Registry</a> link.</p>
+
−
<p>We encourage teams to start completing documentation for their parts on the Registry as soon as you have it available. The sooner you put up your parts, the better you will remember all the details about your parts. Remember, you don't need to send us the DNA sample before you create an entry for a part on the Registry. (However, you <b>do</b> need to send us the DNA sample before the Jamboree. If you don't send us a DNA sample of a part, that part will not be eligible for awards and medal criteria.)</p>
+
  
 +
        <header class="intro">
 +
        <div class="intro-body">
 +
          <div class="container">
 +
            <div class="row">
 +
              <div class="col-md-12">
 +
                <h1 class="brand-heading">Team Parts</h1>
 +
                <p class="intro-text">
 +
                </p>
 +
                <div align="center">
 +
                    <a href="#accordion">
 +
                        <span class="arrowtext">Scroll down to read more</span>
 +
                        <img src="https://static.igem.org/mediawiki/2014/3/3e/Aalto_Helsinki_Nuoli.png" class="arrow">
 +
                    </a>
 +
                </div>
 +
              </div>
 +
            </div>
 +
          </div>
 +
        </div>
 +
      </header>
 +
  <div class="container"> 
 +
    <div class="panel-group" id="accordion" role="tablist" aria-multiselectable="true">
 +
      <div class="panel panel-default">
 +
        <div class="panel-heading" role="tab" id="headingOne">
 +
          <h4 class="panel-title">
 +
            <a class="collapsed" role="button" data-toggle="collapse" data-parent="#accordion" href="#collapseOne" aria-expanded="false" aria-controls="collapseOne">
 +
              Heroin Esterase BBa_K1615045
 +
            </a>
 +
          </h4>
 +
        </div>
 +
        <div id="collapseOne" class="panel-collapse collapse" role="tabpanel" aria-labelledby="headingOne">
 +
          <div class="panel-body">
 +
              <p style="color: black;">
 +
              Heroin esterase, an acetylmorphine carboxylesterase,  was isolated from <i>Rhodococcus erythropolis</i> strain H1 in 1994 from the garden soil at Cambridge and is able to use heroin as its sole carbon and energy source<sup>1</sup>. The gene <i>her</i> encodes this enzyme and has the ability to be expressed in the chassis <i>Escherichia coli</i><sup>2</sup>. The pH optimum for this enzyme to function is in 8.5 in bicine buffer<sup>1</sup>.
 +
<br>
 +
<br>
 +
The activity of heroin esterase can be tested using 4-nitrophenyl acetate which is hydrolised(?) by heroin esterase to form 4-nitrophenol + actetate. This produces a yellow colour as well as being able to be read at 410 nm.
 +
<br>
 +
<br>
 +
Design: The sequence for our enzyme used the original sequence from Rathbone, et al., and was then codon optimised for <i>E. coli</i>. The RFC25 prefix and suffix were added along which required all illegal sites (EcoRI, SpeI, AgeI, NotI, NgoMIV and XbaI) to be removed. As this was a difficult sequence to make as a gBlock, it was ordered as a gene in an ampicillin backbone where it was then digested and ligated into the pSB1C3 backbone.
 +
<br>
 +
<br>
 +
<sup>1</sup>Cameron, G. W., Jordan, K. N., Holt, P. J., Baker, P. B., Lowe, C. R., & Bruce, N. C. (1994). Identification of a heroin esterase in Rhodococcus sp. strain H1. Applied and environmental microbiology, 60(10), 3881-3883.
  
−
<h4>What information do I need to start putting my parts on the Registry?</h4>
+
<br><sup>2</sup>Rathbone, D. A., Holt, P. J., Lowe, C. R., & Bruce, N. C. (1997). Molecular analysis of the Rhodococcus sp. strain H1 her gene and characterization of its product, a heroin esterase, expressed in Escherichia coli. <i>Applied and environmental microbiology</i>, 63(5), 2062-2066.
−
<p>The information needed to initially create a part on the Registry is:</p>
+
−
<ul>
+
−
<li>Part Name</li>
+
−
<li>Part type</li>
+
−
<li>Creator</li>
+
−
<li>Sequence</li>
+
−
<li>Short Description (60 characters on what the DNA does)</li>
+
−
<li>Long Description (Longer description of what the DNA does)</li>
+
−
<li>Design considerations</li>
+
−
</ul>
+
  
−
<p>
+
              </ul>
−
We encourage you to put up <em>much more</em> information as you gather it over the summer. If you have images, plots, characterization data and other information, please also put it up on the part page. </p>
+
            </p>
  
 +
            <div align="center">
 +
                <a href="#" class="btn btn-primary btn-lg outline" role="button">Check it out in the registry</a>
 +
            </div>
 +
         
 +
            </div>
 +
          </div>
 +
        </div>
 +
      </div>
 +
      <div class="panel panel-default">
 +
        <div class="panel-heading" role="tab" id="headingTwo">
 +
          <h4 class="panel-title">
 +
            <a class="collapsed" role="button" data-toggle="collapse" data-parent="#accordion" href="#collapseTwo" aria-expanded="false" aria-controls="collapseTwo">
 +
            Morphine-6-Dehydrogenase BBa_K1615000
 +
            </a>
 +
          </h4>
 +
        </div>
 +
        <div id="collapseTwo" class="panel-collapse collapse" role="tabpanel" aria-labelledby="headingTwo">
 +
          <div class="panel-body">
 +
              <p>
 +
                The structural gene morphine-6-dehyrogenase (<i>morA</i>) was first isolated from <i>Pseudomonas putida</i> M10 as it is capable of growth with morphine as its sole carbon source<sup>1</sup>. Morphine dehydrogenase catalyses the oxidation of both morphine and codeine to produce morphinone and codeinone. During this process NADP<sup>+</sup> is reduced to NADPH which means that it is frequently used to detect morphine and codeine enzymatically<sup>2</sup>.
 +
<br>
 +
<br>
 +
To test the morphine dehydrogenase activity it can be coupled with codeine and NADP<sup>+</sup> to produce codeinone and NADPH. The amount of NADPH produced can be measured at x nm.
 +
<br>
 +
<br>
 +
Design: To make this gene standardised it was codon optomised for the chassis <i>Esherichia coli</i> as well as making it RFC25 compatible which required getting rid of all illegal restriction sites in the gene sequence.
 +
<br>
 +
<br>
 +
<sup>1</sup>Bruce, N. C., Wilmot, C. J., Jordan, K. N., Trebilcock, A. E., Stephens, L. D. G., & Lowe, C. R. (1990). Microbial degradation of the morphine alkaloids: identification of morphinone as an intermediate in the metabolism of morphine by Pseudomonas putida M10. <i>Archives of microbiology</i>, 154(5), 465-470.
 +
<br><sup>2</sup>Rathbone, D. A., Holt, P. J., Lowe, C. R., & Bruce, N. C. (1997). Molecular analysis of the Rhodococcus sp. strain H1 her gene and characterization of its product, a heroin esterase, expressed in Escherichia coli. <i>Applied and environmental microbiology</i>, 63(5), 2062-2066.
 +
            </p>
 +
            <div align="center">
 +
                <a href="#" class="btn btn-primary btn-lg outline" role="button">Check it out in the registry</a>
 +
            </div>
 +
           
 +
          </div>
 +
        </div>
 +
      </div>
 +
      <div class="panel panel-default">
 +
        <div class="panel-heading" role="tab" id="headingThree">
 +
          <h4 class="panel-title">
 +
            <a class="collapsed" role="button" data-toggle="collapse" data-parent="#accordion" href="#collapseThree" aria-expanded="false" aria-controls="collapseThree">
 +
              Monoamine oxidase A BBa_K1615022
 +
            </a>
 +
          </h4>
 +
        </div>
 +
        <div id="collapseThree" class="panel-collapse collapse" role="tabpanel" aria-labelledby="headingThree">
 +
          <div class="panel-body">
 +
              <p>
 +
                Monoamine oxidase A is coded by the gene <i>maoA</i> and is subject to catabolite and ammonium ion repression<sup>1</sup>. Amine oxidases that contain copper/topaquinone (TPQ), like monoamine oxidase A, convert primary amines into their corresponding aldehydes, hydrogen peroxide and ammonia<sup>2</sup>.
 +
<br>
 +
<br>
 +
To test the activity of monoamine oxidase A, tyramine can be used as a substrate and its corresponding aldehyde as well as ammonia and hydrogen peroxide will be produced. When the hydrogen peroxide is coupled with horseradish peroxidase and Amplex red, resorufin, a red colour, will be produced.
 +
<br>
 +
<br>
 +
Design: This monoamine oxidase A sequence was found in <i>Klebsiella pneumoniae</i><sup>3</sup> and was codon optimised for the chassis Escherichia coli as well as made RFC25 compatible with the corresponding prefix and suffix and illegal restriction sites were removed.
 +
<br>
 +
<br>
 +
<sup>1</sup>Oka, M., Murooka, Y., & Harada, T. (1980). Genetic control of tyramine oxidase, which is involved in derepressed synthesis of arylsulfatase in Klebsiella aerogenes. <i>Journal of bacteriology</i>, 143(1), 321-327.
 +
<br><sup>2</sup>McIntire, W. S., & Hartmann, C. (1993). Copper-containing amine oxidases. <i>Principles and applications of quinoproteins</i>, 97-171.
 +
<br><sup>3</sup>Sugino, H., Sasaki, M., Azakami, H., Yamashita, M., & Murooka, Y. (1992). A monoamine-regulated Klebsiella aerogenes operon containing the monoamine oxidase structural gene (maoA) and the maoC gene. <i>Journal of bacteriology</i>, 174(8), 2485-2492.
  
 +
            </p>
 +
            <div align="center">
 +
                <a href="#" class="btn btn-primary btn-lg outline" role="button">Check it out in the registry</a>
 +
            </div>
 +
         
 +
          </div>
 +
        </div>
 +
      </div>
 +
   
  
  
−
 
+
</body>
−
 
+
−
<h4>Inspiration</h4>
+
−
<p>We have a created  a <a href="http://parts.igem.org/Well_Documented_Parts">collection of well documented parts</a> that can help you get started.</p>
+
−
 
+
−
<p> You can also take a look at how other teams have documented their parts in their wiki:</p>
+
−
<ul>
+
−
<li><a href="https://2014.igem.org/Team:MIT/Parts"> 2014 MIT </a></li>
+
−
<li><a href="https://2014.igem.org/Team:Heidelberg/Parts"> 2014 Heidelberg</a></li>
+
−
<li><a href="https://2014.igem.org/Team:Tokyo_Tech/Parts">2014 Tokyo Tech</a></li>
+
−
</ul>
+
−
 
+
−
 
+
−
 
+
−
<h4>Part Table </h4>
+
−
</html>
+
−
<groupparts>iGEM015 Example</groupparts>
+
−
<html>
+
−
 
+
−
 
+
−
 
+
−
</div>
+
 
</html>
 
</html>

Revision as of 22:45, 18 September 2015

Team Parts

Heroin esterase, an acetylmorphine carboxylesterase, was isolated from Rhodococcus erythropolis strain H1 in 1994 from the garden soil at Cambridge and is able to use heroin as its sole carbon and energy source1. The gene her encodes this enzyme and has the ability to be expressed in the chassis Escherichia coli2. The pH optimum for this enzyme to function is in 8.5 in bicine buffer1.

The activity of heroin esterase can be tested using 4-nitrophenyl acetate which is hydrolised(?) by heroin esterase to form 4-nitrophenol + actetate. This produces a yellow colour as well as being able to be read at 410 nm.

Design: The sequence for our enzyme used the original sequence from Rathbone, et al., and was then codon optimised for E. coli. The RFC25 prefix and suffix were added along which required all illegal sites (EcoRI, SpeI, AgeI, NotI, NgoMIV and XbaI) to be removed. As this was a difficult sequence to make as a gBlock, it was ordered as a gene in an ampicillin backbone where it was then digested and ligated into the pSB1C3 backbone.

1Cameron, G. W., Jordan, K. N., Holt, P. J., Baker, P. B., Lowe, C. R., & Bruce, N. C. (1994). Identification of a heroin esterase in Rhodococcus sp. strain H1. Applied and environmental microbiology, 60(10), 3881-3883.
2Rathbone, D. A., Holt, P. J., Lowe, C. R., & Bruce, N. C. (1997). Molecular analysis of the Rhodococcus sp. strain H1 her gene and characterization of its product, a heroin esterase, expressed in Escherichia coli. Applied and environmental microbiology, 63(5), 2062-2066.

The structural gene morphine-6-dehyrogenase (morA) was first isolated from Pseudomonas putida M10 as it is capable of growth with morphine as its sole carbon source1. Morphine dehydrogenase catalyses the oxidation of both morphine and codeine to produce morphinone and codeinone. During this process NADP+ is reduced to NADPH which means that it is frequently used to detect morphine and codeine enzymatically2.

To test the morphine dehydrogenase activity it can be coupled with codeine and NADP+ to produce codeinone and NADPH. The amount of NADPH produced can be measured at x nm.

Design: To make this gene standardised it was codon optomised for the chassis Esherichia coli as well as making it RFC25 compatible which required getting rid of all illegal restriction sites in the gene sequence.

1Bruce, N. C., Wilmot, C. J., Jordan, K. N., Trebilcock, A. E., Stephens, L. D. G., & Lowe, C. R. (1990). Microbial degradation of the morphine alkaloids: identification of morphinone as an intermediate in the metabolism of morphine by Pseudomonas putida M10. Archives of microbiology, 154(5), 465-470.
2Rathbone, D. A., Holt, P. J., Lowe, C. R., & Bruce, N. C. (1997). Molecular analysis of the Rhodococcus sp. strain H1 her gene and characterization of its product, a heroin esterase, expressed in Escherichia coli. Applied and environmental microbiology, 63(5), 2062-2066.

Monoamine oxidase A is coded by the gene maoA and is subject to catabolite and ammonium ion repression1. Amine oxidases that contain copper/topaquinone (TPQ), like monoamine oxidase A, convert primary amines into their corresponding aldehydes, hydrogen peroxide and ammonia2.

To test the activity of monoamine oxidase A, tyramine can be used as a substrate and its corresponding aldehyde as well as ammonia and hydrogen peroxide will be produced. When the hydrogen peroxide is coupled with horseradish peroxidase and Amplex red, resorufin, a red colour, will be produced.

Design: This monoamine oxidase A sequence was found in Klebsiella pneumoniae3 and was codon optimised for the chassis Escherichia coli as well as made RFC25 compatible with the corresponding prefix and suffix and illegal restriction sites were removed.

1Oka, M., Murooka, Y., & Harada, T. (1980). Genetic control of tyramine oxidase, which is involved in derepressed synthesis of arylsulfatase in Klebsiella aerogenes. Journal of bacteriology, 143(1), 321-327.
2McIntire, W. S., & Hartmann, C. (1993). Copper-containing amine oxidases. Principles and applications of quinoproteins, 97-171.
3Sugino, H., Sasaki, M., Azakami, H., Yamashita, M., & Murooka, Y. (1992). A monoamine-regulated Klebsiella aerogenes operon containing the monoamine oxidase structural gene (maoA) and the maoC gene. Journal of bacteriology, 174(8), 2485-2492.