Team:Evry/Interlab Study/tests

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                       <FONT color="white"> <h5>Chassis</h5></font>
                       <FONT color="white"> <h5>Chassis</h5></font>
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                         <a href="#chassis"><img class="img-circle" src="https://static.igem.org/mediawiki/2014/7/7e/Chassis_EVRY.png" height="128" width="128" alt="Conference" border="30px" style="box-shadow: 0 0 8px rgba(0, 0, 0, .8); "/></a>
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                         <font colors ="white"><h5>Transposons</h5></font>
                         <font colors ="white"><h5>Transposons</h5></font>
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                         <FONT color="white"><h5>Target</h5></font>
                         <FONT color="white"><h5>Target</h5></font>
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  <div class="tab-pane active" id="chassis">France</div>
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The goal of the interlab study is to obtain GFP fluorescence data from the iGEM teams all around the world for about twenty constructions. This will permit to observe the result repeatability across technics, strains and teams.
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There were two sub-parts in this study:
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<li>The mandatory one: obtain fluorescence data for three specific genetic devices expressing GFP and compare them. (donner les constructions)
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  <a id="igem-link" target="_blank" href="http://parts.igem.org/Promoters/Catalog/Anderson " title="Go to iGEM site">(J23100 to J23119)</a>.
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<li>The Extra Credit assignments: we choose to study the entire Anderson library of constitutive prokaryotic promoters.
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This library corresponds to 19 constitutive promoters containing some nucleotide mutations on the -35 and the -10, as shown figure x. The idea is to compare the expression strength of promoters according to mutations, from the maximum amount of fluorescence data in order to have significant results.
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    To test the expression of promoters, they are cloned into the PSB1C3 vector followed by the
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    <a id="igem-link" target="_blank" href="http://parts.igem.org/Part:BBa_E0240" title="Go to iGEM site">BBa_E0240</a>, a GFP generator. Constructions are inserted in E. coli DH5 alpha that grown on LB medium added with the selection agent, chloramphenicol. Our team decided to test the fluorescence with a 
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    <a id="igem-link" target="_blank" href="http://www.tecan.com/platform/apps/product/index.asp?MenuID=1813&ID=1918&Menu=1&Item=21.2.10.1.1 " title="Go to iGEM site">TECAN infiniteM200</a> in 96 well plates.
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    Fluorescence data were analyzed via following formula that come from <a id="igem-link" target="_blank" href="http://www.biomedcentral.com/1752-0509/4/55 " title="Go to iGEM site">2010 De Jong et al </a><i> Experimental and computational validation of models of fluorescent and luminescent reporter genes in bacteria </i>.
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    <center><img alt="IMAGE" src="https://static.igem.org/mediawiki/2014/8/88/BMC_formlula.emf.jpg" width="300px;float:left;" class="thumbimage"/><br/><center/><br/>
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  Equation A: A(t) is the corrected absorbance of DH5 alpha with the functional reporter system, Au(t)the uncorrected  absorbance of DH5 alpha with the functional reporter system and Ab(t)the background absorbance corresponding to LB  with chloramphenicol medium absorbance.<br>
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  Equation B: B(t) is the absorbance of the strain carrying the promoterless vector, Iu(t) the uncorrected fluorescence intensity of DH5 alpha with the functional reporter system and Ib(t) the background fluorescence intensity of LB chloramphenicol medium with the DH5 alpha carrying the promoterless vector.<br>
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<h2>Required Devices</h2>
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    <center><img alt="IMAGE" src="https://static.igem.org/mediawiki/2014/7/78/Gaph_final.png" width="700p" class="thumbimage"/><br/><center/><br/>
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The three required constructions shown various fluorescence intensity at OD 600 nm = 0.5, as shown on bar chart on the right. The same trend was observed between OD 600 nm = 0.1 to 0.8(Data shown on
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<a href="https://2014.igem.org/Team:Evry/Interlab_Study/Notebook" >Interlab study notebook</a>). Fluorescence intensity curve profile according to OD 600 nm corresponded to the expected profile for a constitutive promoter (Data shown on
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<a href="https://2014.igem.org/Team:Evry/Interlab_Study/Notebook" >Interlab study notebook</a>).
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<br/><div align="right"><img src="https://static.igem.org/mediawiki/2014/7/7d/Interlab_final.png" width="500px"; alt="image not found" /></div>
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<h4>Entire Anderson library of constitutive promoters (J23100-J23119)</h4>
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  <left> Anderson collection promoter sequences and mutations. </center>
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  <left> Anderson collection promoter sequences and mutations. </center>
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Revision as of 16:10, 17 October 2014

IGEM Evry 2014

Interlab study - Aim & Results

Chassis
 Conference
Transposons
 Workshops
Target
 Hackathons
France
Angleterre
Japon


Contacts


Retrieved from "http://2014.igem.org/Team:Evry/Interlab_Study/tests"