Difference between revisions of "Team:DTU-Denmark/Journal"
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<nav class="navbar navbar-default navbar-fixed-top" role="navigation"> | <nav class="navbar navbar-default navbar-fixed-top" role="navigation"> | ||
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− | <ul class="nav navbar-nav | + | <ul class="nav navbar-nav"> |
− | + | <li class="dropdown"> | |
− | + | <a href="/Team:DTU-Denmark" | |
− | + | >Home | |
− | + | </a></li> | |
− | + | <li class="dropdown"> | |
− | + | <a href="/Team:DTU-Denmark" | |
− | + | class="dropdown-toggle" | |
− | <li> | + | data-toggle="dropdown" |
+ | role="button" | ||
+ | aria-expanded="false">Team and Attributions | ||
+ | <span class="caret"></span></a><ul class="dropdown-menu" role="menu"> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Team" | ||
+ | >Team | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Attributions" | ||
+ | >Attributions | ||
+ | </a></li></ul></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark" | ||
+ | class="dropdown-toggle" | ||
+ | data-toggle="dropdown" | ||
+ | role="button" | ||
+ | aria-expanded="false">Project | ||
+ | <span class="caret"></span></a><ul class="dropdown-menu" role="menu"> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Overview" | ||
+ | >Overview | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Background" | ||
+ | >Background | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/MAGE" | ||
+ | >MAGE Subtilis | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Surfactin" | ||
+ | >Surfactin | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Tyrocidine" | ||
+ | >Tyrocidine | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Chip" | ||
+ | >Lab-on-a-disc | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Intein" | ||
+ | >Intein | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Project/Detection" | ||
+ | >Detection of NRP | ||
+ | </a></li></ul></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Practices" | ||
+ | >Human Practices | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Part_Collection" | ||
+ | >Parts | ||
+ | </a></li> | ||
+ | <li class="dropdown active"> | ||
+ | <a href="/Team:DTU-Denmark/Journal" | ||
+ | >Journal | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Software" | ||
+ | >Software | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark" | ||
+ | class="dropdown-toggle" | ||
+ | data-toggle="dropdown" | ||
+ | role="button" | ||
+ | aria-expanded="false">Achievements | ||
+ | <span class="caret"></span></a><ul class="dropdown-menu" role="menu"> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Achievements" | ||
+ | >Overview of Achievements | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Collaborations" | ||
+ | >Collaborations | ||
+ | </a></li> | ||
+ | <li class="dropdown"> | ||
+ | <a href="/Team:DTU-Denmark/Judging%20Form" | ||
+ | >Judging Form | ||
+ | </a></li></ul></li> | ||
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− | <ul class="nav | + | <ul class="nav navbar-nav"> |
− | <li><a class="page-scroll" href="#Summary">Summary</a></li><li><a class="page-scroll" href="# | + | <li><a class="page-scroll" href="#Summary">Summary</a></li><li><a class="page-scroll" href="#Protocols">Protocols</a></li><li><a class="page-scroll" href="#Timeline">Timeline</a></li><li><a class="page-scroll" href="#References">References</a></li> |
</ul> | </ul> | ||
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− | <div class="jumbotron hero | + | <div class="jumbotron hero bg-hero"> |
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− | <img src=" | + | <img src="/wiki/images/a/af/DTU-Denmark_igemlogo.png"> |
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<div class="row hero-text"> | <div class="row hero-text"> | ||
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− | <a id=" | + | <a id="Protocols-submenu" class="btn btn-default btn-transparent btn-lg page-scroll" href="#Protocols">Protocols</a> |
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− | <a id=" | + | <a id="Timeline-submenu" class="page-scroll" href="#Timeline">Timeline</a> |
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− | <a id=" | + | <a id="References-submenu" class="page-scroll" href="#References">References</a> |
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Scroll down for more<br> | Scroll down for more<br> | ||
− | <a href="#" class="btn btn-circle btn-transparent page-scroll"> | + | <a href="#Summary" class="btn btn-circle btn-transparent page-scroll"> |
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Summary | Summary | ||
</h1> | </h1> | ||
− | <p>For OGRE (Oligo Recombineering) to be effective there is two genetic improvements that can be done. The first and most important is that the organism need to express a recombinase protein such as the beta protein derived from the lambda RED E. coli phage. The second improvement is to destroy/inhibit the mismatch repair system [ | + | <p>For OGRE (Oligo Recombineering) to be effective there is two genetic improvements that can be done. The first and most important is that the organism need to express a recombinase protein such as the beta protein derived from the lambda RED E. coli phage. The second improvement is to destroy/inhibit the mismatch repair system [1]. In B. subtilis the genes mutS and mutL are taking care of the mismatch repair [2]. For proof of concept purposes deleting the mismatch repair system will be adequate. According to (REF) the protein encoded by mutL will not be functional without a functional mutS, this is why deletion of only mutS will be sufficient to take out the mismatch repair system. With respect to recombinase we will test two different recombinases: lambda-beta (codon optimized for <em>B. subtilis 168</em>) and gp35 (a recombinase from the B. subtilis phage SPP1) [3]. For duing proof of concept of OGRE in <em>B. subtilis</em> four different strains was producere via genetic recombineering. All derived from <em>Bacillus subtilis 168</em>:</p> |
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<h1> | <h1> | ||
− | + | Protocols | |
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− | <div aria-multiselectable="true" class="panel-group" id="accordion" role="tablist"> | + | |
<div class="panel panel-default"> | <div class="panel panel-default"> | ||
<div class="panel-heading" id="headingOne" role="tab"> | <div class="panel-heading" id="headingOne" role="tab"> | ||
− | <h4 class="panel-title"><a aria-controls="collapseOne" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseOne" role="button"> | + | <h4 class="panel-title"><a aria-controls="collapseOne" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseOne" role="button">1% Agarose Gel</a></h4> |
</div> | </div> | ||
<div aria-labelledby="headingOne" class="panel-collapse collapse" id="collapseOne" role="tabpanel"> | <div aria-labelledby="headingOne" class="panel-collapse collapse" id="collapseOne" role="tabpanel"> | ||
<div class="panel-body"> | <div class="panel-body"> | ||
− | < | + | <div class="panel-body">Needed materials: 100 ml. TAE buffer. 1gr. agarose. ethidium bromide</div> |
− | < | + | <div class="panel-body">Mix 100 ml. TAE buffer with 1 gr. agarose powder, heat in microwave till the agarose is disolved, let it cool till it can be handled with bare hands, add 10 µl ethidium bromide. pour into mold containing slots for wells, and let cool.</div> |
− | </div> | + | |
</div> | </div> | ||
</div> | </div> | ||
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<div class="panel-heading" id="headingTwo" role="tab"> | <div class="panel-heading" id="headingTwo" role="tab"> | ||
− | <h4 class="panel-title"><a aria-controls="collapseTwo" aria-expanded="false" class="collapsed" data-parent="# | + | <h4 class="panel-title"><a aria-controls="collapseTwo" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseTwo" role="button">Agarose Gel Electrophoresis</a></h4> |
</div> | </div> | ||
<div aria-labelledby="headingTwo" class="panel-collapse collapse" id="collapseTwo" role="tabpanel"> | <div aria-labelledby="headingTwo" class="panel-collapse collapse" id="collapseTwo" role="tabpanel"> | ||
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">Needed materials: 100 ml. TAE buffer. 1gr. agarose. ethidium bromide</div> | ||
− | < | + | <p aria-controls="ui-id-36" aria-expanded="true" aria-selected="false" role="tab" tabindex="0">set gel into Electrophoresis chamber filled with 1X TAE buffer so the wells are covered. <br /> |
+ | mix 5 µl of your PCR reactions/sample with 1 µl 6X loading buffer.<br /> | ||
+ | pipette 5 µl into each well, pour 2-3 µl ladder in some so the size can be seen.<br /> | ||
+ | run at 140 V, for 30-40 min</p> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
− | |||
<div class="panel panel-default"> | <div class="panel panel-default"> | ||
<div class="panel-heading" id="headingThree" role="tab"> | <div class="panel-heading" id="headingThree" role="tab"> | ||
− | <h4 class="panel-title"><a aria-controls="collapseThree" aria-expanded="false" class="collapsed" data-parent="# | + | <h4 class="panel-title"><a aria-controls="collapseThree" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseThree" role="button">Chemocompetent cells</a></h4> |
</div> | </div> | ||
<div aria-labelledby="headingThree" class="panel-collapse collapse" id="collapseThree" role="tabpanel"> | <div aria-labelledby="headingThree" class="panel-collapse collapse" id="collapseThree" role="tabpanel"> | ||
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">Chemo competens is a way of getting E. coli to be able to take up new DNA, since this cant be done naturally like B. subtilis, then chemocompetent cells have to be made.</div> | ||
+ | |||
+ | <div class="panel-body">Protocol can be found <a href="/wiki/images/c/cd/DTU-Denmark_Chemocompetentcells_protocol.pdf" target="_blank">here</a></div> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
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<div class="panel panel-default"> | <div class="panel panel-default"> | ||
<div class="panel-heading" id="headingFour" role="tab"> | <div class="panel-heading" id="headingFour" role="tab"> | ||
− | <h4 class="panel-title"><a aria-controls="collapseFour" aria-expanded="false" class="collapsed" data-parent="# | + | <h4 class="panel-title"><a aria-controls="collapseFour" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseFour" role="button">Glycerol stock</a></h4> |
</div> | </div> | ||
<div aria-labelledby="headingFour" class="panel-collapse collapse" id="collapseFour" role="tabpanel"> | <div aria-labelledby="headingFour" class="panel-collapse collapse" id="collapseFour" role="tabpanel"> | ||
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">Needed materials: a 40% glycerol solution, bacteria of your choice, cryo tubes</div> | ||
+ | |||
+ | <div class="panel-body">mix glycerol and bacteria 50/50 in a cryo tube and place in a -80 frezzer till needed</div> | ||
</div> | </div> | ||
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− | |||
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<div class="panel-heading" id="headingThree" role="tab"> | <div class="panel-heading" id="headingThree" role="tab"> | ||
− | <h4 class="panel-title"><a aria-controls=" | + | <h4 class="panel-title"><a aria-controls="collapseFive" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseFive" role="button">Making plates</a></h4> |
</div> | </div> | ||
− | <div aria-labelledby=" | + | <div aria-labelledby="headingFive" class="panel-collapse collapse" id="collapseFive" role="tabpanel"> |
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <p>A lot of lab work requires plates in order to produce results, and often you need a special concentration, a unique antibiotic mixed into the plates or a different composition than what is available at the lab you are in. If that is the case, the plate needs to be made from scratch, or somewhat along the way.</p> | ||
+ | |||
+ | <p>Protocol can be found <a href="/wiki/images/e/e2/DTU-Denmark_making_plates_protocol.pdf" target="_blank">her</a></p> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
− | |||
<div class="panel panel-default"> | <div class="panel panel-default"> | ||
− | <div class="panel-heading" id=" | + | <div class="panel-heading" id="headingSix" role="tab"> |
− | <h4 class="panel-title"><a aria-controls=" | + | <h4 class="panel-title"><a aria-controls="collapseSix" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseSix" role="button">Minimal media</a></h4> |
</div> | </div> | ||
− | <div aria-labelledby=" | + | <div aria-labelledby="headingSix" class="panel-collapse collapse" id="collapseSix" role="tabpanel"> |
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">For our experiments we have used minimal media designed for B. subtilis. The plates have been made by following this protocol (for 1 liter)</div> | ||
+ | |||
+ | <div class="panel-body">Protocolcan be found <a href="/wiki/images/5/52/DTU-Denmark_Minimal_media_protocol.pdf" target="_blank">here</a></div> | ||
</div> | </div> | ||
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− | <div class="panel-heading" id=" | + | <div class="panel-heading" id="headingSeven" role="tab"> |
− | <h4 class="panel-title"><a aria-controls=" | + | <h4 class="panel-title"><a aria-controls="collapseSeven" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseSeven" role="button">Miniprep</a></h4> |
</div> | </div> | ||
− | <div aria-labelledby=" | + | <div aria-labelledby="headingSeven" class="panel-collapse collapse" id="collapseSeven" role="tabpanel"> |
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">in order to work with plasmids, then they needs to be purified, and condensed, this is done bu minipreping the bacteria, and afterwards the concentration can be investigating using NanoDrop™.</div> | ||
+ | |||
+ | <div class="panel-body">Protocol <a href="/wiki/images/4/48/DTU-Denmark_Miniprep_protocol.pdf" target="_blank">here</a>.</div> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
− | |||
<div class="panel panel-default"> | <div class="panel panel-default"> | ||
− | <div class="panel-heading" id=" | + | <div class="panel-heading" id="headingEight" role="tab"> |
− | <h4 class="panel-title"><a aria-controls=" | + | <h4 class="panel-title"><a aria-controls="collapseEigth" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseEigth" role="button">Natural competent Bacillus subtilis 168</a></h4> |
</div> | </div> | ||
− | <div aria-labelledby=" | + | <div aria-labelledby="headingEigth" class="panel-collapse collapse" id="collapseEigth" role="tabpanel"> |
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">Bacillus can be made Competent which makes it easy to alter the genome, so wanted mutaions can be inserted, this is a unique way of investigateing new pathways.</div> | ||
+ | |||
+ | <div class="panel-body">Protocol can be found <a href="/wiki/images/2/29/DTU-Denmark_natural_competent_subtilis_protocol.pdf" target="_blank">here</a></div> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
− | |||
<div class="panel panel-default"> | <div class="panel panel-default"> | ||
− | <div class="panel-heading" id=" | + | <div class="panel-heading" id="headingNine" role="tab"> |
− | <h4 class="panel-title"><a aria-controls=" | + | <h4 class="panel-title"><a aria-controls="collapseNine" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseNine" role="button">OGRE</a></h4> |
</div> | </div> | ||
− | <div aria-labelledby=" | + | <div aria-labelledby="headingNine" class="panel-collapse collapse" id="collapseNine" role="tabpanel"> |
− | <div class="panel-body"> | + | <div class="panel-body"> |
+ | <div class="panel-body">Electrotoration of bacteria is used in order to introduce mutations in the lagging strand of the strain, we have chosen to name this OGRE, this technique are used in order to incorporate wanted mutations in a strain of Bacillus subtilis 168.</div> | ||
+ | |||
+ | <div class="panel-body">Protocol can be found <a href="/wiki/images/0/02/DTU-Denmark_OGRE_protocol.pdf" target="_blank">here</a></div> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
− | |||
<div class="panel panel-default"> | <div class="panel panel-default"> | ||
<div class="panel-heading" id="headingTen" role="tab"> | <div class="panel-heading" id="headingTen" role="tab"> | ||
− | <h4 class="panel-title"><a aria-controls=" | + | <h4 class="panel-title"><a aria-controls="collapseTen" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseTen" role="button">Preperation of Electro Competent Bacillus subtilis 168</a></h4> |
</div> | </div> | ||
<div aria-labelledby="headingTen" class="panel-collapse collapse" id="collapseTen" role="tabpanel"> | <div aria-labelledby="headingTen" class="panel-collapse collapse" id="collapseTen" role="tabpanel"> | ||
+ | <div class="panel-body"> | ||
+ | <div class="panel-body">Electro competent cells are used in our OGRE part of the project, so a batch of cells was prepared using the following protocol.</div> | ||
+ | |||
+ | <div class="panel-body">Protocol can be found <a href="/wiki/images/2/2e/DTU-Denmark_Electro_competent_protocol.pdf" target="_blank">here</a></div> | ||
+ | </div> | ||
+ | </div> | ||
+ | </div> | ||
+ | |||
+ | <div class="panel panel-default"> | ||
+ | <div class="panel-heading" id="headingEleven" role="tab"> | ||
+ | <h4 class="panel-title"><a aria-controls="collapseEleven" aria-expanded="false" class="collapsed" data-parent="#accordion" data-toggle="collapse" href="#collapseEleven" role="button">Collapsible Group Item #11</a></h4> | ||
+ | </div> | ||
+ | |||
+ | <div aria-labelledby="headingTen" class="panel-collapse collapse" id="collapseEleven" role="tabpanel"> | ||
<div class="panel-body">000.</div> | <div class="panel-body">000.</div> | ||
</div> | </div> | ||
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</div> | </div> | ||
</div> | </div> | ||
− | + | ||
− | + | ||
<div id="Timeline"> | <div id="Timeline"> | ||
+ | <div class="container"> | ||
+ | <div class="row"> | ||
+ | <div class="col-lg-12"> | ||
+ | <h2>Timeline</h2> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | <ul class="timeline"> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Biology of the Future </h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-09-14</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p style="text-align: justify;"><img alt="" src="/wiki/images/c/c5/DTU-Denmark_timeline_novoamb.png" style="height: 158px; width: 250px; float: left;" />Our team member Pernille spent an afternoon giving a presentation on cell factories and synthetic biology to 30 highschool students. She says: "Synthetic biology is all about being creative and use your imagination to design bio-solutions that can impact the world. For that exact reason, it is neccesary to discuss which ethical values tat should drive research". The highschool students had lots of questions and opinion for the discussion about ethics.</p> | ||
+ | |||
+ | <p style="text-align: justify;"> </p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">VWR to the rescue! </h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-09-01</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that the VWR is contributing our team with their awesome products!</p> | ||
+ | |||
+ | <p><img alt="" src="None" style="width: 400px; height: 533px;" /></p> | ||
+ | |||
+ | <p> </p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Visit from Fisher Scientific! </h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-26</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>Today we had a visit by Susanne Basse from Fisher Scientific. She came by our office with a wagon full of items, Thanks! </p> | ||
+ | |||
+ | <p><img alt="" src="/wiki/images/c/c7/DTU-Denmark_Fisher_Scientific_visit.png" style="width: 300px; height: 280px;" /></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contribution from Macrogen!</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-20</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are pleased to announce that Macrogen is sponsoring our team with sequencing!</p> | ||
+ | |||
+ | <p><a href="http://www.macrogen.com/" target="_blank"><img alt="" src="/wiki/images/1/11/DTU-Denmark_Marcogen.jpg" style="width: 300px; height: 199px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contributions from AKG!</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-19</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that AKG Acoustics is sponsoring our team with headphones for presentations in Boston! </p> | ||
+ | |||
+ | <p><a href="http://us.akg.com/akg-homepage-us.html" target="_blank"><img alt="" src="/wiki/images/9/9d/DTU-Denmark_AKG.png" style="width: 400px; height: 121px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contributions from VWR - Bie & Berntsen A/S</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-11</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that VWR - Bie & Berntsen A/S is sponsoring our team with lab consumeables!</p> | ||
+ | |||
+ | <p><a href="dk.vwr.com" target="_blank"><img alt="" src="/wiki/images/e/ea/DTU-Denmark_VWR.jpg" style="width: 400px; height: 76px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contributions from Fisher Scientific</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-11</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that Fisher Scientific is sponsoring our team with lab materials! </p> | ||
+ | |||
+ | <p><a href="https://dk.fishersci.com/dk/" target="_blank"><img alt="" src="/wiki/images/b/b4/DTU-Denmark_Fisher_Scientific.jpg" style="width: 400px; height: 123px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Canoeing</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-07</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>Today we took a day off from lab to go canoeing, followed by a barbeque at Chris' place.</p> | ||
+ | |||
+ | <p><img alt="" src="/wiki/images/6/6a/DTU-Denmark_Teambuilding.jpg" /></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contribution from SnapGene</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-08-03</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that the SnapGene is contributing our team with lisences to their awesome product!</p> | ||
+ | |||
+ | <p><a href="http://www.snapgene.com/" target="_blank"><img alt="" src="/wiki/images/5/58/DTU-Denmark_Snapgene.png" style="width: 400px; height: 113px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Wiki Wizard</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-07-30</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We have uploaded the first beta version of the Wiki Wizard to<a href="https://github.com/dkmva/igem-wiki-wizard/commits/master"> <span class="fa fa-github " style="color:#000000;"></span>GitHub</a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">First BioBrick cloned into B. subtilis</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-07-30</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>An expression cassette with lambda beta recombinase was sucesfully transformed into <em>Bacillus subtilis</em>. First step towards establishing MAGE in <em>Bacillus subtilis</em> completed!</p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">BioBrick High School Project</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-07-23</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>3 high school students; Simran, Noor, and Charlotte, joined us and Biotech Academy for 5 days to make biosensors that changes color, when water is poluted. Read more at <span style="color:#000000;"><span style="background-color:#FFFF00;">ADD</span></span> and <a href="http://biotechacademy.dk">http://biotechacademy.dk</a>.</p> | ||
+ | |||
+ | <p style="text-align: center;"><img alt="" src="/wiki/images/b/b8/DTU-Denmark_highschool_biotechacademy.jpg" style="width: 350px; height: 197px;" /></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">First sequencing results</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-07-22</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We received our first sequencing results. One step closer to our first BioBrick!</p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Brevibacillus parabrevis arrives</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-07-16</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p><em>Brevibacillus parabrevis </em>produces a lot of different antibiotics including tyrocidine. We will transfer the tyrocidine operon to our chassis <em>Bacillus subtilis</em>.</p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contribution from the Lundbeck Foundation!</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-07-15</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that the Lundbeck Foundation is contributing financially to our team!</p> | ||
+ | |||
+ | <p><a href="http://www.lundbeckfoundation.com/" target="_blank"><img alt="" src="/wiki/images/e/e2/DTU-Denmark_Lundbeckfonden.jpg" style="width: 348px; height: 96px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contributions from In Vitro!</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-05-06</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that In Vitro A/S is sponsoring our team with lab consumeables! </p> | ||
+ | |||
+ | <p><a href="in-vitro.dk" target="_top"><img alt="" src="/wiki/images/7/7c/DTU-Denmark_In_Vitro.jpg" style="width: 400px; height: 156px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contributions from Frisnette</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-04-30</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that Frisnette is sponsoring our team with lab consumeables! </p> | ||
+ | |||
+ | <p><a href="http://frisenette.dk/" target="_blank"><img alt="" src="/wiki/images/c/c8/DTU-Denmark_Frisenette.jpg" style="width: 330px; height: 85px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">We get our strain</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-04-28</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <div class="pull-left"><img alt="" src="/wiki/images/e/e1/DTU-Denmark_timeline_W168.jpg" style="width: 120px; height: 160px;" /></div> | ||
+ | |||
+ | <div class="pull-left"> | ||
+ | <p>We get a plate of our chassis strain <em>Bacillus subtilis</em> W168.<br /> | ||
+ | We can now begin the work in the laboratory.</p> | ||
+ | </div> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">BioBrick Workshop</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-04-24</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p style="text-align: center;"><img alt="" src="/wiki/images/f/fe/DTU-Denmark_timeline_biobrickworkshop.jpg" style="width: 350px; height: 198px;" /></p> | ||
+ | |||
+ | <p>Southern University of Denmark and University of Copenhagen joined us for a three-day BioBrick tutorial, where we spent three days in the laboratory cloning, learning about iGEM, wiki design, and getting to know each other through social activities.</p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Contribution from the Otto Mønsted Fonden!</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-03-08</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>We are happy to announce that the Otto Mønsted Fonden is contributing financially to our team!</p> | ||
+ | |||
+ | <p><a href="http://www.ottomoensted.dk/" target="_blank"><img alt="" src="/wiki/images/3/35/DTU-Denmark_Ottomoensted.png" style="width: 400px; height: 85px;" /></a></p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">First official team meeting</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2015-01-14</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>Today we had our first official team meeting. We talked about the project and got to know each other.</p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | <li class="timeline-inverted"> | ||
+ | <div class="timeline-badge"><i class="glyphicon glyphicon-check"></i></div> | ||
+ | <div class="timeline-panel"> | ||
+ | <div class="timeline-heading"> | ||
+ | <h4 class="timeline-title">Introductory meeting</h4> | ||
+ | <p><small class="text-muted"><i class="glyphicon glyphicon-time"></i> 2014-11-25</small></p> | ||
+ | </div> | ||
+ | <div class="timeline-body"> | ||
+ | <p>This day, last years team held an introductory session about iGEM for interested people.</p> | ||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </li> | ||
+ | |||
+ | </ul> | ||
+ | </div> | ||
+ | </div> | ||
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+ | |||
+ | <div id="References"> | ||
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</h1> | </h1> | ||
− | + | <ol> | |
+ | |||
+ | <li>Carr, P. A., Wang, H. H., Sterling, B., Isaacs, F. J., Lajoie, M. J., Xu, G., … Jacobson, J. M. (2012). Enhanced multiplex genome engineering through co-operative oligonucleotide co-selection. Nucleic Acids Research, 40(17). doi:10.1093/nar/gks455</li> | ||
+ | |||
+ | <li>Ginetti, F., Perego, M., Albertini, A. M., & Galizzi, A. (1996). Bacillus subtilis mutS mutL operon: Identification, nucleotide sequence and mutagenesis. Microbiology, 142(8), 2021–2029. doi:10.1099/13500872-142-8-2021</li> | ||
+ | |||
+ | <li>Sun, Z., Deng, A., Hu, T., Wu, J., Sun, Q., Bai, H., … Wen, T. (2015). A high-efficiency recombineering system with PCR-based ssDNA in Bacillus subtilis mediated by the native phage recombinase GP35. Applied Microbiology and Biotechnology, 99(12), 5151–5162. <a href="http://dx.doi.org/10.1007/s00253-015-6485-5" target="_blank">doi:10.1007/s00253-015-6485-5</a></li> | ||
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Revision as of 22:38, 18 September 2015
Summary
For OGRE (Oligo Recombineering) to be effective there is two genetic improvements that can be done. The first and most important is that the organism need to express a recombinase protein such as the beta protein derived from the lambda RED E. coli phage. The second improvement is to destroy/inhibit the mismatch repair system [1]. In B. subtilis the genes mutS and mutL are taking care of the mismatch repair [2]. For proof of concept purposes deleting the mismatch repair system will be adequate. According to (REF) the protein encoded by mutL will not be functional without a functional mutS, this is why deletion of only mutS will be sufficient to take out the mismatch repair system. With respect to recombinase we will test two different recombinases: lambda-beta (codon optimized for B. subtilis 168) and gp35 (a recombinase from the B. subtilis phage SPP1) [3]. For duing proof of concept of OGRE in B. subtilis four different strains was producere via genetic recombineering. All derived from Bacillus subtilis 168:
- ∆amyE::beta-neoR
- ∆amyE::GP35-neoR
- ∆mutS::beta-neoR
- ∆mutS::GP35-neoR
The four different strains where compared to each other to get compare there transformation efficienc in MAGE.
Protocols
set gel into Electrophoresis chamber filled with 1X TAE buffer so the wells are covered.
mix 5 µl of your PCR reactions/sample with 1 µl 6X loading buffer.
pipette 5 µl into each well, pour 2-3 µl ladder in some so the size can be seen.
run at 140 V, for 30-40 min
A lot of lab work requires plates in order to produce results, and often you need a special concentration, a unique antibiotic mixed into the plates or a different composition than what is available at the lab you are in. If that is the case, the plate needs to be made from scratch, or somewhat along the way.
Protocol can be found her
Timeline
-
Biology of the Future
2015-09-14
Our team member Pernille spent an afternoon giving a presentation on cell factories and synthetic biology to 30 highschool students. She says: "Synthetic biology is all about being creative and use your imagination to design bio-solutions that can impact the world. For that exact reason, it is neccesary to discuss which ethical values tat should drive research". The highschool students had lots of questions and opinion for the discussion about ethics.
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VWR to the rescue!
2015-09-01
We are happy to announce that the VWR is contributing our team with their awesome products!
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Visit from Fisher Scientific!
2015-08-26
Today we had a visit by Susanne Basse from Fisher Scientific. She came by our office with a wagon full of items, Thanks!
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-
-
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Canoeing
2015-08-07
Today we took a day off from lab to go canoeing, followed by a barbeque at Chris' place.
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Wiki Wizard
2015-07-30
We have uploaded the first beta version of the Wiki Wizard to GitHub
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First BioBrick cloned into B. subtilis
2015-07-30
An expression cassette with lambda beta recombinase was sucesfully transformed into Bacillus subtilis. First step towards establishing MAGE in Bacillus subtilis completed!
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BioBrick High School Project
2015-07-23
3 high school students; Simran, Noor, and Charlotte, joined us and Biotech Academy for 5 days to make biosensors that changes color, when water is poluted. Read more at ADD and http://biotechacademy.dk.
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First sequencing results
2015-07-22
We received our first sequencing results. One step closer to our first BioBrick!
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Brevibacillus parabrevis arrives
2015-07-16
Brevibacillus parabrevis produces a lot of different antibiotics including tyrocidine. We will transfer the tyrocidine operon to our chassis Bacillus subtilis.
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-
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We get our strain
2015-04-28
We get a plate of our chassis strain Bacillus subtilis W168.
We can now begin the work in the laboratory. -
BioBrick Workshop
2015-04-24
Southern University of Denmark and University of Copenhagen joined us for a three-day BioBrick tutorial, where we spent three days in the laboratory cloning, learning about iGEM, wiki design, and getting to know each other through social activities.
-
-
First official team meeting
2015-01-14
Today we had our first official team meeting. We talked about the project and got to know each other.
-
Introductory meeting
2014-11-25
This day, last years team held an introductory session about iGEM for interested people.
References
- Carr, P. A., Wang, H. H., Sterling, B., Isaacs, F. J., Lajoie, M. J., Xu, G., … Jacobson, J. M. (2012). Enhanced multiplex genome engineering through co-operative oligonucleotide co-selection. Nucleic Acids Research, 40(17). doi:10.1093/nar/gks455
- Ginetti, F., Perego, M., Albertini, A. M., & Galizzi, A. (1996). Bacillus subtilis mutS mutL operon: Identification, nucleotide sequence and mutagenesis. Microbiology, 142(8), 2021–2029. doi:10.1099/13500872-142-8-2021
- Sun, Z., Deng, A., Hu, T., Wu, J., Sun, Q., Bai, H., … Wen, T. (2015). A high-efficiency recombineering system with PCR-based ssDNA in Bacillus subtilis mediated by the native phage recombinase GP35. Applied Microbiology and Biotechnology, 99(12), 5151–5162. doi:10.1007/s00253-015-6485-5
Department of Systems Biology
Søltofts Plads 221
2800 Kgs. Lyngby
Denmark
P: +45 45 25 25 25
M: dtu-igem-2015@googlegroups.com