Team:DTU-Denmark-2

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<b>Plug'n'play with DNA</b><br><br>
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<b>Making Molecular Biology Easier</b>
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<br>
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<b>We have designed a novel standardized assembly system, called “Plug 'n' Play with DNA” where ready to use biological parts can be gathered without the use of restriction enzymes and ligase. This will make cloning faster and assembly of expression vectors possible within a few hours. We have created a library of standardized biological parts for mammalian cells and <i>Aspergilli </i> ready to plug 'n' play. You can find more information about how this standard works <a href="https://2011.igem.org/Team:DTU-Denmark-2/Project/PlugnplayAssembly">here</a>.<b>
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<b>A Reporter System</b><br><br>
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The emerging and fast growing field of molecular and synthetic biology calls for simpler, faster and more efficient cloning techniques. Conventional cloning techniques include restriction digestion and ligation, which can be both time consuming and complex – especially when working with mammalian cells and fungi.
 
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So in order to optimize the cloning process to become faster and more efficient, it would be convenient to avoid the use of restriction enzymes.
 
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In early 1990s, the uracil excision-based cloning was invented as a ligation-independent cloning technique that could substitute the conventional cloning with restriction enzymes and ligase. This technique was further developed, and a few years later, New England Biolabs (NEB) introduced the USERTM  Friendly Cloning Kit, which was to ensure cloning without the use of restriction enzymes. However, the kit was not compatible with proofreading polymerases, since they stalled when encountering a uracil base in the DNA template, as it is a promutanegic event (Nour-Eldin et al., 2010; New England Biolabs, 2004).
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However, newer proofreading polymerases have been developed that can read through the uracils and thus be compatible with the concept of USER. This has opened the door to a more simple and flexible approach to molecular cloning techniques.
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The method applies long complementary overhangs on the PCR product as well as the destination vector. These overhangs can anneal each other to form a stable hybridization product. The overhangs on the PCR product are custom made and independent of restriction sites, which is very convenient when working with fungi and mammalian cells.
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We will develop a standardized cloning system, called “Plug’n’Play”, where certain categories of biological parts can be gathered. This means that the parts in the form of pre-produced PCR-products are directly mixed with a vector, which makes assembly of an expression vector possible within a few hours. All the parts in the form of PCR-products will be distributed in microtiter plates directly ready for cloning. Furthermore, the “Plug’n’Play” kit will contain a back-up where all parts are contained on a plasmid to ensure amplification of a mutation free template if needed. 
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The simple and easy use of the system will be demonstrated by developing a reporter targeting system for the fungus Aspergillus niger. Furthermore, will we demonstrate its application in mammalian cells."</p>
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As proof of concept we have created a reporter system that can be used for anything from monitoring gene expression to determination of protein localization. We expressed and localized fluorescent proteins in the model organism <a href="https://2011.igem.org/Team:DTU-Denmark-2/results/Proofofconcept/fungi" ><i>Aspergillus nidulans</i></a> and the <a href="https://2011.igem.org/Team:DTU-Denmark-2/results/Proofofconcept/mammalian"> mammalian cell line U-2 OS </a> with great success.</p>
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<b>Flexibility</b><br><br>
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<img src="https://static.igem.org/mediawiki/2011/a/aa/Plasmid_puzzle_colour.jpg" width="187px" height="50px"> </img><br>
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Standardization entails rigidity. Therefore we have written a <a href="https://2011.igem.org/Team:DTU-Denmark-2/Project/PlugnplayAssembly/customization" >guide</a> that allows the researcher to customize the system, so proteins can be assembled seamless, multiple mutations can be introduced in one round of cloning etc. The only limitation to this is your creativity.</p>
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<b>Applications</b><br><br>
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<a href="https://2011.igem.org/Team:DTU-Denmark-2/Project/introduction#Application%20area">Here</a> you can learn more about the numerous applications of the Plug’n’Play with DNA assembly standard and the research areas, where this system is especially advantageous.</p>
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<b>Data Page</b><br><br>
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You can find an overview of our project on our <a href="https://2011.igem.org/Team:DTU-Denmark-2/results/data_page">Data Page</a>. Here you also find our favorite submitted biobricks, and the biobricks we characterized during the summer.</p>
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<b>Achievements</b><br><br>
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<img src="https://static.igem.org/mediawiki/2011/f/f7/Achievements.jpg" width="187px" height="50px"> </img><br>
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Designing a novel assembly standard and creating a reporter system for <i>Aspergillus nidulans</i> and mammalian cells are our main achievements. You can read more about our accomplishments on <a href="https://2011.igem.org/Team:DTU-Denmark-2/Project/achievements"> this page</a>.</p>
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<b>The Team</b><br><br>
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We are five girls that have been working on this project for three months with support from our three supervisors. You can learn more about us, and how we started an iGEM team on our <a href="https://2011.igem.org/Team:DTU-Denmark-2/Team">Team Page</a>.</p>
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<a href="https://igem.org/Team_Wikis?year=2011"><b>iGEM 2011 Wiki Main Page</b></a>
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  slogan[0] = "It’s fast, it’s furious, it’s Plug’n’Play with DNA";
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  slogan[1] = "Pack your ligases and restriction enzymes away Plug’n’play with DNA is here to stay";
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  slogan[2] = "Take action and improve your reaction";
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  slogan[3] = "Making research easy and without sorrow, our Plug’n’play kit you can borrow";
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  slogan[4] = "Making research easy and without tears, with our Plug’n’play kit you can save years";
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  slogan[5] = "Dry your tears and save some years Plug’n’play Is here to stay";
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  slogan[6] = "Use Plug’n’play and reasearch will be a joy every day";
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  slogan[7] = "Plug’n’play with DNA, an easy way to save your day";
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  slogan[8] = "Plug’n’play with DNA, it’s like Speedy Gonzales in microtiter plates";
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  slogan[9] = "Plug’n’play with DNA will change the world today";
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  slogan[10] = "Plug’n’play can hold your favorite DNA";
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  slogan[11] = "Mix it, heat it, use it, it's that easy";
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  slogan[12] = "Avoid restriction, use the plug 'n' play edition";
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  slogan[13] = "Make a smart decision, use the plug 'n' play edition";
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  slogan[14] = "Plug’n’play with DNA is complete, it is easy, cool and smart, indeed";
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<b> Sponsored by</b><br><br>
<b> Sponsored by</b><br><br>
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<img src="https://static.igem.org/mediawiki/2011/a/a2/Novozymes.jpeg" with="150px"height="150px"> </img>&nbsp;
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<img src="https://static.igem.org/mediawiki/2010/4/47/Lundbeckfonden.gif"height="40px"> </img> <br><br>
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<img src="https://static.igem.org/mediawiki/2011/0/0c/Alk_Logo.jpg" with="80px"height="80px"> </img> &nbsp;
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<img src="https://static.igem.org/mediawiki/2011/7/74/Novo_nordisk_logo.jpg" with="150px"height="150px"> </img> &nbsp;
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<img src="https://static.igem.org/mediawiki/2011/2/21/IDTLogo2010.png"with="40px"height="40px"> </img> &nbsp;
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<img src="https://static.igem.org/mediawiki/2011/9/90/Pharmadanmark.png"with="50px"height="50px"> </img> &nbsp;
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<img src="https://static.igem.org/mediawiki/2011/2/22/Dtu-uk-a2-200.png" with="50px"height="50px" > </img> &nbsp;
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Latest revision as of 23:10, 21 September 2011


Making Molecular Biology Easier


We have designed a novel standardized assembly system, called “Plug 'n' Play with DNA” where ready to use biological parts can be gathered without the use of restriction enzymes and ligase. This will make cloning faster and assembly of expression vectors possible within a few hours. We have created a library of standardized biological parts for mammalian cells and Aspergilli ready to plug 'n' play. You can find more information about how this standard works here.


A Reporter System

As proof of concept we have created a reporter system that can be used for anything from monitoring gene expression to determination of protein localization. We expressed and localized fluorescent proteins in the model organism Aspergillus nidulans and the mammalian cell line U-2 OS with great success.
Flexibility

Standardization entails rigidity. Therefore we have written a guide that allows the researcher to customize the system, so proteins can be assembled seamless, multiple mutations can be introduced in one round of cloning etc. The only limitation to this is your creativity.
Applications

Here you can learn more about the numerous applications of the Plug’n’Play with DNA assembly standard and the research areas, where this system is especially advantageous.


Data Page

You can find an overview of our project on our Data Page. Here you also find our favorite submitted biobricks, and the biobricks we characterized during the summer.
Achievements

Designing a novel assembly standard and creating a reporter system for Aspergillus nidulans and mammalian cells are our main achievements. You can read more about our accomplishments on this page.
The Team

We are five girls that have been working on this project for three months with support from our three supervisors. You can learn more about us, and how we started an iGEM team on our Team Page.

iGEM 2011 Wiki Main Page









Sponsored by

  

 

   

       

Comments or questions to the team? Please

Retrieved from "http://2011.igem.org/Team:DTU-Denmark-2"