Difference between revisions of "Team:RHIT/Parts"

 
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<h1>Parts</h1>
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<p> To see more information about our parts, including some design considerations and the sources of our parts, please check out the pages on the registry. </p>
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<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>
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<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>
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<h3>Note</h3>
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<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>
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<h3>Adding parts to the registry</h3>
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<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>
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<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>
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<a href="http://parts.igem.org/Add_a_Part_to_the_Registry">
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ADD PARTS
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<h3>Inspiration</h3>
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<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>
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<p> You can also take a look at how other teams have documented their parts in their wiki:</p>
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<h4> Plasmid 1 Parts </h4>
 
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<li><a href="https://2014.igem.org/Team:MIT/Parts"> 2014 MIT </a></li>
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<li> <p style="font-weight:bold">BBa_K2716000 - Optimized Double-Mutant PETase </p>PETase is an aromatic polyesterase that breaks PET into MHET, as well as some terephthalic acid and BHET. The double-mutated optimized PETase contains a S238F/W159H mutation, allowing for a more efficient enzyme. <br><br></li>
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<li><a href="https://2014.igem.org/Team:Heidelberg/Parts"> 2014 Heidelberg</a></li>
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<li> <p style="font-weight:bold">BBa_K2716002 - MHETase </p> MHETase breaks MHET into terephthalic acid and ethylene glycol.<br><br> </li>
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<li><a href="https://2014.igem.org/Team:Tokyo_Tech/Parts">2014 Tokyo Tech</a></li>
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<li> <p style="font-weight:bold">BBa_K2716100 - Complete Plasmid 1 </p>This composite part contains a promoter, a ribosomal binding site, the double-mutated optimized PETase, a secretion tag (pelB), MHETase, and a terminator. PETase is an aromatic polyesterase that breaks PET into MHET, as well as some terephthalic acid and BHET. The double-mutated optimized PETase contains a S238F/W159H mutation, allowing for a more efficient enzyme. PelB is a secretion tag that will allow the PETase enzyme to be secreted outside of the cell. This increases the efficiency of the PET breakdown. MHETase breaks MHET into terephthalic acid and ethylene glycol. The second secretion tag is added so that the MHETase enzyme will also be secreted. <br><br></li>
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<li> <p style="font-weight:bold">BBa_K2716101 - Promoter+RBS+pelB+PETase </p> This composite part contains a promoter, a ribosomal binding site, the double-mutated optimized PETase, and a secretion tag (pelB). PETase is an aromatic polyesterase that breaks PET into MHET, as well as some terephthalic acid and BHET. The double-mutated optimized PETase contains a S238F/W159H mutation, allowing for a more efficient enzyme. PelB is a secretion tag that will allow the PETase enzyme to be secreted outside of the cell. This increases the efficiency of the PET breakdown. This composite is designed for the testing of PETase in a vector that does not previously contain a promoter nor a terminator. Therefore this would cause an overexpression of PETase, theoretically overstressing the cell to death. I.E. it's a killswitch.<br><br></li>
 
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<h4> Plasmid 2 </h4>
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<li> <p style="font-weight:bold"> BBa_K2716004 - Promoter </p> LacI-repressed pTrc promoter <br><br></li>
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<li> <p style="font-weight:bold"> BBa_K2716102 - Complete Plasmid 2</p> This composite part contains the sequence that outlines the metabolic cycle that turns ethylene glycol into malate, which is then used by the cell as a carbon source. This is through glycolaldehyde reductase turning ethylene glycol into glycoladehyde. Glycoladehyde dehydrogenase then turns this compound into glycolate. Glycolate oxidase turns glycolate into glyoxylate, which is then turned into malate by malate synthase. <br><br> </li>
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<li> <p style="font-weight:bold"> BBa_K2716200 - Glycolate Oxidase Subunit D </p> The first of three subunits that make up glycolate oxidase. Glycolate oxidase transforms glycolate into glyoxylate. <br><br> </li>
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<li><p style="font-weight:bold"> BBa_K2716201 - Glycolate Oxidase Subunit E</p> The second of three subunits that make up glycolate oxidase. Glycolate oxidase transforms glycolate into glyoxylate. <br><br> </li>
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<li> <p style="font-weight:bold"> BBa_K2716202 - Glycolate Oxidase Subunit F </p> The last of three subunits that make up glycolate oxidase. Glycolate oxidase transforms glycolate into glyoxylate. <br><br> </li>
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<li> <p style="font-weight:bold"> BBa_K2716203 - Glycolate Oxidase Composite </p> This is a composite of three subunits which form glycolate oxidase when expressed. Glycolate oxidase transforms glycolate into glyoxylate. <br><br>
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<p> The Glycolate oxidase we submitted was cloned from /E. coli/ MG1655 and contains three subunits. It was inserted into the pETDuet vector for expression and was induced by 0.1 and 0.2 mM IPTG at 30ºC for 20 hours.
  
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All three subunit of Glycolate Oxidase(GO) are shown in the SDS-PAGE. GO contains three subunits, D,E, and F, which weigh 55.5 kDa, 38.3 kDa, and 45.0 kDa respectively. Lane 1 was BL21(DE3) with pETDuet-GO, but without IPTG induction process. Lanes 2 and 3 were BL21(DE3) with empty pETDute vectors and contain 0.1 and 0.2 mM IPTG. Lanes 4 to 7 are BL21(De3) with pETDuet-GO. Lanes 4 and 6 were induced with 0.1mM IPTG, and lanes 5 and 7 were induced with 0.2mM IPTG. All the inductions were done at 30ºC.
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<img style="width:70%" src = "https://static.igem.org/mediawiki/2018/a/a6/T--RHIT--SDSpage.jpg"></center></li>
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<li> <p style="font-weight:bold"> BBa_K2716003 - Terminator used in both plasmids </p> Terminator for E. coli pSB1C3 backbone used in BBa_K2716100 and BBa_K2716102 composite parts. <br><br> </li>
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<h3>What information do I need to start putting my parts on the Registry?</h3>
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<p>The information needed to initially create a part on the Registry is:</p>
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<li>Part Name</li>
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<li>Part type</li>
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<li>Creator</li>
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<li>Sequence</li>
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<li>Short Description (60 characters on what the DNA does)</li>
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<li>Long Description (Longer description of what the DNA does)</li>
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<li>Design considerations</li>
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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>
 
  
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<h3>Part Table </h3>
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<p>Please include a table of all the parts your team has made during your project on this page. Remember part characterization and measurement data must go on your team part pages on the Registry. </p>
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<groupparts>iGEM18 RHIT</groupparts>
 
<groupparts>iGEM18 RHIT</groupparts>
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Latest revision as of 21:03, 17 October 2018




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patekm@rose-hulman.edu
5500 Wabash Ave.
Terre Haute, IN 47803

To see more information about our parts, including some design considerations and the sources of our parts, please check out the pages on the registry.

Plasmid 1 Parts

  • BBa_K2716000 - Optimized Double-Mutant PETase

    PETase is an aromatic polyesterase that breaks PET into MHET, as well as some terephthalic acid and BHET. The double-mutated optimized PETase contains a S238F/W159H mutation, allowing for a more efficient enzyme.

  • BBa_K2716002 - MHETase

    MHETase breaks MHET into terephthalic acid and ethylene glycol.

  • BBa_K2716100 - Complete Plasmid 1

    This composite part contains a promoter, a ribosomal binding site, the double-mutated optimized PETase, a secretion tag (pelB), MHETase, and a terminator. PETase is an aromatic polyesterase that breaks PET into MHET, as well as some terephthalic acid and BHET. The double-mutated optimized PETase contains a S238F/W159H mutation, allowing for a more efficient enzyme. PelB is a secretion tag that will allow the PETase enzyme to be secreted outside of the cell. This increases the efficiency of the PET breakdown. MHETase breaks MHET into terephthalic acid and ethylene glycol. The second secretion tag is added so that the MHETase enzyme will also be secreted.

  • BBa_K2716101 - Promoter+RBS+pelB+PETase

    This composite part contains a promoter, a ribosomal binding site, the double-mutated optimized PETase, and a secretion tag (pelB). PETase is an aromatic polyesterase that breaks PET into MHET, as well as some terephthalic acid and BHET. The double-mutated optimized PETase contains a S238F/W159H mutation, allowing for a more efficient enzyme. PelB is a secretion tag that will allow the PETase enzyme to be secreted outside of the cell. This increases the efficiency of the PET breakdown. This composite is designed for the testing of PETase in a vector that does not previously contain a promoter nor a terminator. Therefore this would cause an overexpression of PETase, theoretically overstressing the cell to death. I.E. it's a killswitch.

Plasmid 2

  • BBa_K2716004 - Promoter

    LacI-repressed pTrc promoter

  • BBa_K2716102 - Complete Plasmid 2

    This composite part contains the sequence that outlines the metabolic cycle that turns ethylene glycol into malate, which is then used by the cell as a carbon source. This is through glycolaldehyde reductase turning ethylene glycol into glycoladehyde. Glycoladehyde dehydrogenase then turns this compound into glycolate. Glycolate oxidase turns glycolate into glyoxylate, which is then turned into malate by malate synthase.

  • BBa_K2716200 - Glycolate Oxidase Subunit D

    The first of three subunits that make up glycolate oxidase. Glycolate oxidase transforms glycolate into glyoxylate.

  • BBa_K2716201 - Glycolate Oxidase Subunit E

    The second of three subunits that make up glycolate oxidase. Glycolate oxidase transforms glycolate into glyoxylate.

  • BBa_K2716202 - Glycolate Oxidase Subunit F

    The last of three subunits that make up glycolate oxidase. Glycolate oxidase transforms glycolate into glyoxylate.

  • BBa_K2716203 - Glycolate Oxidase Composite

    This is a composite of three subunits which form glycolate oxidase when expressed. Glycolate oxidase transforms glycolate into glyoxylate.

    The Glycolate oxidase we submitted was cloned from /E. coli/ MG1655 and contains three subunits. It was inserted into the pETDuet vector for expression and was induced by 0.1 and 0.2 mM IPTG at 30ºC for 20 hours. All three subunit of Glycolate Oxidase(GO) are shown in the SDS-PAGE. GO contains three subunits, D,E, and F, which weigh 55.5 kDa, 38.3 kDa, and 45.0 kDa respectively. Lane 1 was BL21(DE3) with pETDuet-GO, but without IPTG induction process. Lanes 2 and 3 were BL21(DE3) with empty pETDute vectors and contain 0.1 and 0.2 mM IPTG. Lanes 4 to 7 are BL21(De3) with pETDuet-GO. Lanes 4 and 6 were induced with 0.1mM IPTG, and lanes 5 and 7 were induced with 0.2mM IPTG. All the inductions were done at 30ºC.

  • BBa_K2716003 - Terminator used in both plasmids

    Terminator for E. coli pSB1C3 backbone used in BBa_K2716100 and BBa_K2716102 composite parts.

<groupparts>iGEM18 RHIT</groupparts>