Team:Genspace/parts

From 2014.igem.org

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<h2>What is the Open Lab?</h2>
<h2>What is the Open Lab?</h2>
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<p>Genspace has a unique opportunity to expand its impact beyond its physical presence in New York and bring hands-on involvement with synthetic biology to schools and communities around the world. The Open Lab is the key step towards this vision; it is the complete set of knowledge, tools, and resources required to successfully develop a thriving community biolab, all created following open source principles. Enthusiast communities and organizations around the world can partner with the Open Lab to fill in key gaps in knowledge, tools, or resources to launch their own community biolabs.</p>
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<div id='part_status_wrapper'><div style='display:block;position:relative;height:16px;padding-top:3px;text-align:center;border:1px solid gray;;background-color:#ffffff;'><a href='http://parts.igem.org/Help:Part_Status_Box'>Not Released</a></div>
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<div class='sample_results' style='display:block;position:relative;height:16px;padding-top:3px;text-align:center;border:1px solid gray;border-top:none;;background-color:#ffffff';'><a href='http://parts.igem.org/Help:Part_Status_Box'>Sample No part sequence</a></div>
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<h2>What is currently being developed for the Open Lab?</h2>
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<div style='display:block;position:relative;height:16px;padding-top:3px;text-align:center;border:1px solid gray;border-top:none;'background-color:white;'>No Results</div>
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<div style='display:block;position:relative;height:16px;padding-top:3px;text-align:center;border:1px solid gray;border-top:none;'><a href='http://parts.igem.org/partsdb/uses.cgi?part=BBa_K1429002'>-1 Uses</a></div>
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<p>Genspace is currently focusing on two essentials projects that will form the core of the knowledge and tool components for the Open Lab.</p>
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<div style='display:block;position:relative;height:16px;padding-top:3px;text-align:center;border:1px solid gray;border-top:none; border:none;'><a href='http://parts.igem.org/partsdb/get_part.cgi?part=BBa_K1429002''> Get This Part</A></div></div><div style='float:right;position:relative;'><img style='padding-top:0px' src='http://parts.igem.org/images/partbypart/icon_translational_unit.png'>
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              <div style='display:block;font-size:90%;line-height:90%' title='Part Type'>Translational_Unit</div>
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<p><strong>Open Lab Blueprint</strong> – visit <a alt="Open Lab Blueprint" href="OpenLab-Blueprint.org" target="_blank">OpenLab-Blueprint.org</a> to view
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              <div style='display:block;position:relative;top:3px;font-size:90%;line-height:100%' type='Nickname'></div>
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              </div>
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<p>The Open Lab Blueprint is an easy-to-follow guide to launch and develop a sustainable community biolab. The website is a work in progress and is hosting at OpenLab-Blueprint.org. This Blueprint is a holistic view of the community biolab and includes the following components:</p>
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      <h1 class='firstHeading'>Part:BBa_K1429002</h1>  <div style= 'position: relative;font-size:95%;line-height:100%;'>
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                Designed by: Christal Gordon &nbsp;  Group: iGEM14_Genspace  &nbsp; (2014-09-29)</div>
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<ul>
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              <div style='height:1em;'></div> <div id="bodyContent">
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<li>Starting up a lab, including identifying the right space, acquisition of key equipment, and efficiently implementing safety standards and best practices.</li>
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<div id="contentSub"></div>
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<li>Developing a community, including building awareness, developing programs of engagement for students, the amateur scientists, and the general public, and collaborating with the broader scientific and educational communities.</li>
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<!--
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<li>Creating unique content, including educational resources, using the Genspace curriculum and engaging activities, focused on tailored, site-specific explorations and experiments.</li>
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<div id="jump-to-nav">Jump to:                        <a href="#column-one">navigation</a>, <a href="#searchInput">search</a></div>-->
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<li>Operating a biolab, including a resource base of protocols for standard genetic engineering activities, geared towards a budget-constrained lab, operational best-practices ranging from procurement of reagents to proper storage of experiments, and financial sustainability models.</li>
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<!-- start content -->
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<p><SPAN style='font-size: 150%; font-weight: bold;'>Cyan Fluorescent Protein (CFP) "Cindy Lou" coding region, intellectual property-free</SPAN>
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<p>The Open Plasmid is a key tool for the community biolab. Genspace has partnered with Cathal Garvey, founder of Indie Biotech, to design an open source and IP-free plasmid for the community biolab. The Open Plasmid is a core platform on which many experiments in a community biolab can be built on, essentially the equivalent of an Arduino for the citizen science community. The plasmid is designed to be easily modifiable, and therefore approachable to everyone from weekend researcher to curious high school student. The plasmid is also designed to be easy to cultivate with minimal resources. Unlike most plasmids, it does not need to be grown with antibiotics, significantly increasing accessibility to budget-constrained organizations, and having less negative environmental impact.</p>
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</p><p>IP free cyan fluorescent protein. We have a ribosome binding site (RBS).
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</p>
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<p>As a first step, we have cloned the IP-free fluorescent protein genes. These are meant to be used together with the Open Plasmid. Each gene codes for a differently fluorescent protein: cyan, green, and red. These sequences can be used as teaching tools for classes or workshops or as molecular tools by independent researchers who are part of the community.</p>
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<h4> <span class="mw-headline" id="Background">Background</span></h4>
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<div class="entry-results"><p>We want to take Tuesday&#39;s PCR products and put them into the pSB1C3 backbone.&nbsp;</p>
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<p><strong>Digest PCRs:</strong></p>
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<p>10 ul PCR product</p>
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<p>2 ul cutsmart buffer (10x stock)</p>
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<p>1 ul PstI</p>
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<p><u>1 ul EcoRI</u></p>
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<p>20 ul total --&gt; incubate for 30 min at 37C</p>
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<p>&nbsp;</p>
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<p><strong>PCR purify digest product (only 14 ul - save 6 ul):</strong></p>
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<p>Follow kit protocol. Elute in elution buffer.</p>
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<p>Worried that the washed columns won&#39;t bind DNA, we are going to use some of the set-aside (unpurified) digest product for a backup ligation. We&#39;ll run a gel of our purification, but we are going to set up a ligation beforehand, so we won&#39;t have even rough estimates of DNA concentrations.</p>
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<p>Set up ligations:</p>
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<table style="width: 500px;">
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&lt;tbody&gt;
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<tr>
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<td><strong>Component</strong></td>
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<td><strong>Using purified digest product</strong></td>
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<td><strong>Using unpurified digest product</strong></td>
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<td><strong>BB alone</strong></td>
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</tr>
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<tr>
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<td>dH2O</td>
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<td>x</td>
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<td>11</td>
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<td>14</td>
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</tr>
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<tr>
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<td>Insert (RFP or GFP)</td>
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<td>14</td>
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<td>3</td>
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<td>x</td>
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</tr>
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<tr>
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<td>1:10 BB</td>
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<td>3</td>
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<td>3</td>
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<td>3</td>
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</tr>
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<tr>
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<td>T4 buffer (10</td>
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<td>1</td>
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<td>1</td>
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<td>1</td>
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</tr>
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<tr>
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<td>&nbsp;</td>
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<td>&nbsp;</td>
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<td>&nbsp;</td>
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<td>&nbsp;</td>
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</tr>
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&lt;/tbody&gt;
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</table>
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<p>The above were incubated 30 min at RT then stored at -20C.</p>
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<p>&nbsp;</p>
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</div>
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<p><br />
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&lt;/div&gt;
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</p>
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<div class="entry">
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<h4> <span class="mw-headline" id="Results">Results</span></h4>
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<div class="entry-results"><p>We ran a 1% gel of the digest before and after purification. We had a decent yield, maybe 40% of our initial digest product in the purified lanes.&nbsp;</p>
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</div>
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<p><br />
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</p>
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</div>
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<div class="entry">
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<h4> <span class="mw-headline" id="Conclusions">Conclusions</span></h4>
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<div class="entry-results"><p>Next step: Transform the ligated plasmids into E. coli!</p>
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</div>
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<p><br />
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</p>
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</div>
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<p><br />

Latest revision as of 00:46, 8 October 2014

Project

What is the Open Lab?

Translational_Unit

Part:BBa_K1429002

Designed by: Christal Gordon   Group: iGEM14_Genspace   (2014-09-29)

Cyan Fluorescent Protein (CFP) "Cindy Lou" coding region, intellectual property-free

IP free cyan fluorescent protein. We have a ribosome binding site (RBS).

Background

We want to take Tuesday's PCR products and put them into the pSB1C3 backbone. 

Digest PCRs:

10 ul PCR product

2 ul cutsmart buffer (10x stock)

1 ul PstI

1 ul EcoRI

20 ul total --> incubate for 30 min at 37C

 

PCR purify digest product (only 14 ul - save 6 ul):

Follow kit protocol. Elute in elution buffer.

Worried that the washed columns won't bind DNA, we are going to use some of the set-aside (unpurified) digest product for a backup ligation. We'll run a gel of our purification, but we are going to set up a ligation beforehand, so we won't have even rough estimates of DNA concentrations.

Set up ligations:

<tbody> </tbody>
Component Using purified digest product Using unpurified digest product BB alone
dH2O x 11 14
Insert (RFP or GFP) 14 3 x
1:10 BB 3 3 3
T4 buffer (10 1 1 1
       

The above were incubated 30 min at RT then stored at -20C.

 


</div>

Results

We ran a 1% gel of the digest before and after purification. We had a decent yield, maybe 40% of our initial digest product in the purified lanes. 


Conclusions

Next step: Transform the ligated plasmids into E. coli!



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