Difference between revisions of "Team:ETH Zurich/Materials"

 
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<h1>Materials</h1>
 
<h1>Materials</h1>
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<h2>Bacterial strains and Mammalian Cell Lines</h2>
 
<h2>Bacterial strains and Mammalian Cell Lines</h2>
  
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<h3>TOP10</h3>
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<h3><i>Escherichia coli</i> TOP10</h3>
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<p>The bacterial strain we used as a base for all our transformations was TOP10.</p>
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<p>The bacterial strain we used as a base for all our transformations was TOP10 (Genotype: F- mcrA &Delta;(mrr-hsdRMS-mcrBC) &phi;80lacZ&Delta;M15 &Delta;lacX74 nupG recA1 araD139 &Delta;(ara-leu)7697 galE15 galK16 rpsL(StrR) endA1 &lambda;-). These cells were provided by the Panke Group at ETH Zurich.</p>
  
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<p>A list of all strains we generated in the course of our project can be found here (link).</p>
 
  
 
<h3>Jurkat Cells</h3>
 
<h3>Jurkat Cells</h3>
  
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<p>Immortalized human T lymphocyte cell line. We used this cell line as a model line for cancer cells with elevated lactate production and TRAIL susceptibility (papers).</p>
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<p>Immortalized human T lymphocyte cell line. We used this cell line as a model line for cancer cells with elevated lactate production and TRAIL susceptibility (papers). A big thank you to the Tay Group at ETH Zurich who kindly offered us an aliquot of their cells.</p>
  
 
<h3>HL60</h3>
 
<h3>HL60</h3>
  
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<p>Cell line derived from acute myeloid leukemia. We used this cell line as a model line for cancer cells with elevated lactate production and TRAIL susceptibility (papers).</p>
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<p>Cell line derived from acute myeloid leukemia. We used this cell line as a model line for cancer cells with elevated lactate production and TRAIL susceptibility (papers). Many thanks to the Neri Group at ETH Zurich who kindly offered us an aliquot of their cells.</p>
  
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<h3>3T3</h3>
 
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<p>Murine fibroblast cell line. We used this cell line as a model line for healthy cells which are restistant to TRAIL and produce normal levels of lactate.</p>
 
  
 
<h3>SK-BR-3</h3>
 
<h3>SK-BR-3</h3>
  
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<p>HER2-positive breast cancer cell line. We used this cell line to validate the chimeric receptors of the <a href="https://2015.igem.org/Team:Stockholm">Stockholm</a> iGEM team.</p>
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<p>HER2-positive breast cancer cell line. We used this cell line to validate the chimeric receptors of the <a href="https://2015.igem.org/Team:Stockholm">Stockholm</a> iGEM team. We are very grateful to Michal Stanczak from the University Hospital of Basel for offering us a sample of this cell line.</p>
  
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<h3>3T3</h3>
  
 +
<p>Murine fibroblast cell line. We used this cell line as a model line for healthy cells which are restistant to TRAIL and produce normal levels of lactate. Many thanks to the Tay Group at ETH Zurich who kindly offered us an aliquot of their cells.</p>
  
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<h3>3T3 P65 -/- </h3>
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<p>Murine fluorescent fibroblast cell line.
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<h2>Machines</h2>
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<div class="info">
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<h3>Nano Drop</h3>
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<ul>
 
<ul>
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<li>Thermo Scientific NanoDrop® 2000 UV-Vis spectrophotometer combined with NanoDrop 2000 / 2000c Software</li>
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<li> H2B-GFP: nuclear staining</li>
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<a href="http://www.nanodrop.com/Productnd2000overview.aspx"><p>for further information click here</p></a>
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<li> P65- dsRed: cytoskeleton</li>
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[[File:NanoDrop.JPG|center|thumb]]
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</ul>
 
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We used this cell line to study the co-culture of the bacteria and the chip. Many thanks to the Tay Group at ETH Zurich who kindly offered us an aliquot of their cells.</p>  
 
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<div class="info">
 
  
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<h3>Plate Reader</h3>
 
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<ul>
 
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<li>Tecan Infinite M200 Pro™ combined with Tecan iControl TM software</li>
 
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<a href="http://www.tecan.com/platform/apps/product/index.asp?MenuID=1812&ID=1916&Menu=1&Item=21.2.10.1"><p>for further information (platereader) click here</p></a>
 
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<a href="http://www.tecan.ch/platform/apps/product/index.asp?MenuID=1817&ID=1924&Menu=1&Item=21.7.8"><p>for further information (software) click here</p></a>
 
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[[File:TecanPlateReader.JPG|center|thumb]]
 
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[[File:ProgramTecanReader.JPG|center|thumb]]
 
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<h2>Buffers and media </h2>
  
 
<div class="info">
 
<div class="info">
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<h3>Annexin V binding buffer</h3>
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<h3>Absorbance Measurments</h3>
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<ul>
 
<ul>
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<li>Novaspec Plus™ Visible Spectrophotometer</li>
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<li>10 mM HEPES</li>
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<a href="http://www.gelifesciences.com/webapp/wcs/stores/servlet/ProductDisplay?categoryId=11012&catalogId=10101&productId=20430&storeId=11782&langId=-1"><p>for further information click here</p></a>
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<li>140 mM NaCl</li>
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<li>2.5 mM</li>
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[[File:ODPhotometer.JPG|center|thumb]]
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<li>250 mM CaCl<sub>2</sub></li>
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<li>pH 7.4</li>
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<li>Filter the solution through a 0.2 &micro;m filter. </li>
 
</ul>
 
</ul>
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</div>
 
  
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<div class="info">
 
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<h3>UV- and Dark Hood</h3>
 
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<ul>
 
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<li>Biostep Dark-Hood DH-50™ combined with Argus-X1™ software and Biostep argusX1™ basic licence</li>
 
−
<a href="http://www.biostep.de/i18n_en_DE/products/Bio_Imaging_components_1776/Tripod__dark_hoods_and_accessories_1863/Dark_hoods_1865/Dark_Hood_DH_40_with_control_panel_and_display_6/index.html"><p>for further information (hood) click here</p></a>
 
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<a href="https://www.biostep.de/products/Bio_Imaging_software_1843/Acquisition_and_control_software_1844/argus_X1___for_cameras_and_scanners_1845/biostep_argusX1__basic_licence_1/index.html"><p>for further information (software) click here</p></a>
 
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[[File:Darkhood.JPG|center|thumb]]
 
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<div class="info">
 
<div class="info">
  
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<h3>Large Centrifuge</h3>
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<h3>Jurkat medium</h3>
 
<ul>
 
<ul>
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<li>Eppendorf Centrifuge 5810 R</li>
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<li>2 mL MEM 1x</li>
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<a href="http://www.eppendorf.com/int/index.php?pb=01edea0fe7a0e358&action=products&contentid=1&catalognode=10019"><p>for further information click here</p></a>
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<li>20 mL FBS 100%</li>
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<li>2 mL GlutaMax 100x</li>
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[[File:BigCentrifuge.JPG|center|thumb]]
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<li>0.4 mL Penicillin (10k units/mL) /Streptomyocin (10k &micro;g/mL )</li>
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<li>200 mL RPMI</li>
 
</ul>
 
</ul>
 
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</div>
 
</div>
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<div class="info">
 
<div class="info">
  
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<h3>Small Centrifuge</h3>
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<h3>Terrific Broth</h3>
 
<ul>
 
<ul>
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<li>Eppendorf Centrifuge 5424 R</li>
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<li>2.32 g/L KH<sub>2</sub>PO<sub>4</sub></li>
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<a href="http://www.labdepotinc.com/Product_Details~id~450~pid~59833.aspx"><p>for further information click here</p></a>
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<li>12.54 g/L K<sub>2</sub>HPO<sub>4</sub></li>
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<li>1.2% Tryptone</li>
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[[File:EppiCentrifuge.JPG|center|thumb]]
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<li>2.4% Yeast Extract</li>
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<li>0.4% Gylcerol</li>
 +
<li>Filter the solution through a 0.2 &micro;m filter.</li>
 
</ul>
 
</ul>
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</div>
 
  
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<div class="info">
 
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<h3>Shaker</h3>
 
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<ul>
 
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<li>Kuhner Climo-Shaker ISF1-X</li>
 
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<a href="http://www.kuhner.com/de/produkte/schuettelmaschinen/inkubatoren/isf1-x.html"><p>for further information click here</p></a>
 
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[[File:ShakerIncubator.JPG|center|thumb]]
 
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</div>
 
</div>
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<div class="info">
 
<div class="info">
  
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<h3>PCR Machine</h3>
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<h3>SOC medium </h3>
 
<ul>
 
<ul>
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<li>Applied Biosystems Veriti 96Well Thermal Cycler</li>
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<li>20 g/L Tryptophane</li>
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<a href="https://www.thermofisher.com/ch/en/home/life-science/pcr/thermal-cyclers-realtime-instruments/thermal-cyclers/veriti-thermal-cycler.html"><p>for further information click here</p></a>
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<li>5 g/L Yeast extract</li>
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<li>0.5 g/L NaCl </li>
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[[File:PCRMachine.JPG|center|thumb]]
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<li>250 mM KCl</li>
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<html>
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<li>1 M MgCl<sub>2</sub></li>
 +
<li>50&#37; (w/v) sterile glucose</li>
 +
<li>Filter the solution through a 0.2 &micro;m filter. </li>
 
</ul>
 
</ul>
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</div>
 
  
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<div class="info">
 
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<h3>Gel Electrophoresis</h3>
 
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<ul>
 
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<li>Helixx Technologies MyRun®</li>
 
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<a href="http://www.helixxtec.com/helixx/c/iMyRun/iMyRun.htm"><p>for further information click here</p></a>
 
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[[File:GelRunningThing.JPG|center|thumb]]
 
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</ul>
 
 
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 
</div>
 
</div>
−
 
  
  
 
<div class="info">
 
<div class="info">
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<h3>TFB1</h3>
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<h3>Eppi-shaker/Heatingblock</h3>
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−
<ul>
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<li>Eppendorf Thermomixer compact</li>
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<a href="http://www.eppendorf.com/int/index.php?pb=93a83e2a7b7b1b05&action=products&contentid=1&catalognode=9840"><p>for further information click here</p></a>
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</html>
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[[File:HeatingBlock.JPG|center|thumb]]
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<html>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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<div class="info">
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−
 
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<h3>Vortex</h3>
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<ul>
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<li>Scientific Industries VortexGenie®2</li>
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<a href="http://www.scientificindustries.com/genie2.html"><p>for further information click here</p></a>
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</html>
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[[File:Vortex.JPG|center|thumb]]
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<html>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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<div class="info">
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<h3>Balance</h3>
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<ul>
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<li>MettlerToledo XS4002S DeltaRange® Balance</li>
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<a href="http://us.mt.com/us/en/home/products/Laboratory_Weighing_Solutions/Precision_Balances/XS_Precision_Balances/XS_4002_S_Dual_Range.html"><p>for further information click here</p></a>
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</html>
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[[File:Balance.JPG|center|thumb]]
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<html>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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<div class="info">
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<h3>Incubator</h3>
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<ul>
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<li>Thermo Scientific Heraeus Incubator</li>
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<a href="https://webshop.fishersci.com/insight2_uk/getProduct.do?productCode=11626250&resultSetPosition=0"><p>for further information click here</p></a>
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</html>
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[[File:Incubator.JPG|center|thumb]]
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<html>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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<div class="info">
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<h3>FACS</h3>
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<ul>
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<li>BD LSRFortessa™ cell analyzer</li>
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<a href="http://www.bdbiosciences.com/us/instruments/research/cell-analyzers/bd-lsrfortessa/m/751752?cc=US"><p>for further information click here</p></a>
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</html>
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[[File:FACS.JPG|center|thumb]]
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<html>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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<div class="info">
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−
<h3>Fluorescence Scanner for Western Blots</h3>
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<ul>
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<li>ODYSSEY® CLx Infrared Imaging System</li>
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<a href="http://www.licor.com/bio/products/imaging_systems/odyssey/"><p>for further information click here</p></a>
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</html>
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[[File:FluorescenceScanner.JPG|center|thumb]]
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<html>
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</ul>
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</div>
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<div class="info">
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<h3>Emulsifier</h3>
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<ul>
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<li>EmulsiFlex-C3 from AVESTIN, Inc. and AVESTIN Europe GmbH/li>
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<a href="http://www.avestin.com/gc3page.html"><p>for further information click here</p></a>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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</div>
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<div class="expContainer">
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<h2>Buffers and media </h2>
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<div class="info">
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<h3>TFP1</h3>
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<ul>
 
<ul>
 
<li>100 mM RbCl</li>
 
<li>100 mM RbCl</li>
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<li>30 mM potassium acetate</li>
 
<li>30 mM potassium acetate</li>
 
<li>10 mM CaCl<sub>2</sub></li>
 
<li>10 mM CaCl<sub>2</sub></li>
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<li>15&#37;glycerol</li>
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<li>15&#37; glycerol</li>
 
</ul>
 
</ul>
 
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 
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<h3>TFP2</h3>
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<h3>TFB2</h3>
 
<ul>
 
<ul>
 
<li>100 mM MOPS</li>
 
<li>100 mM MOPS</li>
 
<li>50 mM RbCl</li>
 
<li>50 mM RbCl</li>
 
<li>75 mM CaCl<sub>2</sub></li>
 
<li>75 mM CaCl<sub>2</sub></li>
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<li>15&#37;glycerol</li>
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<li>15&#37; glycerol</li>
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</ul>
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
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</div>
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<div class="info">
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<h3>SOC medium </h3>
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<ul>
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<li>20 g/L Tryptophane</li>
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<li>5 g/L Yeast extract</li>
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−
<li>0.5 g/L NaCl </li>
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−
<li>250 mM KCl</li>
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−
<li>1M MgCl<sub>2</sub></li>
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−
<li>50&#37; (w/v) sterile glucose</li>
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<li>Filter the solution through a 0.2 &micro;m filter. </li>
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</ul>
 
</ul>
  
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</div>
 
</div>
  
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<div class="info">
 
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<h3>Jurkat medium</h3>
 
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<ul>
 
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<li>2 mL MEM 1x</li>
 
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<li>20 mL FBS 100%</li>
 
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<li>2 mL GlutaMax 100x</li>
 
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<li>0.4 mL Penicillin (10k units/mL) /Streptomyocin (10k &micro;g/mL )</li>
 
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<li>200 mL RPMI</li>
 
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</ul>
 
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 
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</div>
 
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<div class="info">
 
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<h3>Annexin V binding buffer</h3>
 
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<ul>
 
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<li>10 mM HEPES</li>
 
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<li>140 mM NaCl</li>
 
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<li>2.5 mM</li>
 
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<li>250 mM CaCl<sub>2</sub></li>
 
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<li>pH 7.4</li>
 
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<li>Filter the solution through a 0.2 &micro;m filter. </li>
 
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</ul>
 
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 
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<h3>BsaI-HF</h3>
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<ul>
 +
    <li>Product: BsaI-HF® </li>
 +
    <li>Company: New England BioLabs</li>
 +
    <li>REF: R3535S</li>
 +
    <a href="https://www.neb.com/products/r3535-bsai-hf"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
  
  
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</div>
 
</div>
  
 +
<div class="info">
 +
 +
<h3>Protein G coated Beads</h3>
 +
<ul>
 +
    <li>Product: Pierce™ Protein G Magnetic Beads</li>
 +
    <li>Company: Thermo Fischer Scientific</li>
 +
    <li>REF: 88847</li>
 +
    <a href="https://www.thermofisher.com/order/catalog/product/88847"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
  
 
<div class="info">
 
<div class="info">
Line 587: Line 401:
 
     <li>Product: Roti®-MagBeads Streptavidin</li>
 
     <li>Product: Roti®-MagBeads Streptavidin</li>
 
     <li>Company: Carl Roth</li>
 
     <li>Company: Carl Roth</li>
−
     <li>REF: </li>
+
     <li>REF: HP57.2</li>
 
     <a href="https://www.carlroth.com/de/de/Chemikalien/A-Z-Chemikalien/R/Roti%3Csup%3E%C2%AE%3C-sup%3E-MagBeads/Roti%3Csup%3E%C2%AE%3C-sup%3E-MagBeads-Streptavidin/p/0000000a0002a23a00010023_de"><p>for further information click here</p></a>
 
     <a href="https://www.carlroth.com/de/de/Chemikalien/A-Z-Chemikalien/R/Roti%3Csup%3E%C2%AE%3C-sup%3E-MagBeads/Roti%3Csup%3E%C2%AE%3C-sup%3E-MagBeads-Streptavidin/p/0000000a0002a23a00010023_de"><p>for further information click here</p></a>
 
</ul>
 
</ul>
Line 692: Line 506:
 
     <li>REF: 02860-2.5L</li>
 
     <li>REF: 02860-2.5L</li>
 
     <a href="http://www.sigmaaldrich.com/catalog/product/fluka/02860?lang=de&region=CH"><p>for further information click here</p></a>
 
     <a href="http://www.sigmaaldrich.com/catalog/product/fluka/02860?lang=de&region=CH"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>DMEM</h3>
 +
<ul>
 +
    <li>Product: DMEM, high glucose, no glutamine, no phenol red</li>
 +
    <li>Company: Gibco ThermoFisher Scientific</li>
 +
    <li>REF: 31053-028</li>
 +
    <a href="https://www.thermofisher.com/order/catalog/product/31053028"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
 +
<div class="info">
 +
 +
<h3>DMEM with Glutamine</h3>
 +
<ul>
 +
    <li>Product: DMEM cell culture media</li>
 +
    <li>Company: Gibco ThermoFisher Scientific</li>
 +
    <li>REF: 41965-039</li>
 +
    <a href="http://wiki.toxbank.net/wiki/DMEM_cell_culture_media_%2841965-039%29"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
 +
<div class="info">
 +
 +
<h3>RPMI</h3>
 +
<ul>
 +
    <li>Product: RPMI 1640 Medium, no phenol red</li>
 +
    <li>Company: Gibco ThermoFisher Scientific</li>
 +
    <li>REF: 11835-063</li>
 +
    <a href="https://www.thermofisher.com/order/catalog/product/11835063"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
 +
<div class="info">
 +
 +
<h3>Accutase</h3>
 +
<ul>
 +
    <li>Product: StemPro® Accutase® Cell Dissociation Reagent</li>
 +
    <li>Company: Gibco ThermoFisher Scientific</li>
 +
    <li>REF: A11105-01</li>
 +
    <a href="https://www.thermofisher.com/order/catalog/product/A1110501"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>SuperKillerTRAIL™</h3>
 +
<ul>
 +
    <li>Product: SuperKillerTRAIL™ </li>
 +
    <li>Company: Enzo</li>
 +
    <li>REF: N3200SALX-201-115-C010</li>
 +
    <a href="http://www.enzolifesciences.com/ALX-201-115/superkillertrail-soluble-human-recombinant/"><p>for further information click here</p></a>
 +
</ul>
 +
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
<div class="info">
 +
 +
<h3>TRAIL</h3>
 +
<ul>
 +
    <li>Product: TRAIL / APO2 Ligand Human Recombinant </li>
 +
    <li>Company: ProspecBio</li>
 +
    <li>REF: CYT-443</li>
 +
    <a href="http://www.prospecbio.com/APO2L_Human_3_9/"><p>for further information click here</p></a>
 
</ul>
 
</ul>
  
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<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 
</div>
 
</div>
 +
 +
</div>
 +
 +
<div class="expContainer">
 +
 +
<h2>Machines</h2>
 +
 +
<div class="info">
 +
 +
<h3>Absorbance Measurments</h3>
 +
<ul>
 +
<li>Novaspec Plus™ Visible Spectrophotometer</li>
 +
<a href="http://www.gelifesciences.com/webapp/wcs/stores/servlet/ProductDisplay?categoryId=11012&catalogId=10101&productId=20430&storeId=11782&langId=-1"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Balance</h3>
 +
<ul>
 +
<li>MettlerToledo XS4002S DeltaRange® Balance</li>
 +
<a href="http://us.mt.com/us/en/home/products/Laboratory_Weighing_Solutions/Precision_Balances/XS_Precision_Balances/XS_4002_S_Dual_Range.html"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Gel Electrophoresis</h3>
 +
<ul>
 +
<li>Helixx Technologies MyRun®</li>
 +
<a href="http://www.helixxtec.com/helixx/c/iMyRun/iMyRun.htm"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Emulsifier</h3>
 +
<ul>
 +
<li>EmulsiFlex-C3 from AVESTIN, Inc. and AVESTIN Europe GmbH/li>
 +
<a href="http://www.avestin.com/gc3page.html"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Eppi-shaker/Heatingblock</h3>
 +
<ul>
 +
<li>Eppendorf Thermomixer compact</li>
 +
<a href="http://www.eppendorf.com/int/index.php?pb=93a83e2a7b7b1b05&action=products&contentid=1&catalognode=9840"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>FACS</h3>
 +
<ul>
 +
<li>BD LSRFortessa™ cell analyzer</li>
 +
<a href="http://www.bdbiosciences.com/us/instruments/research/cell-analyzers/bd-lsrfortessa/m/751752?cc=US"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
 +
 +
 +
<div class="info">
 +
 +
<h3>Incubator</h3>
 +
<ul>
 +
<li>Thermo Scientific Heraeus Incubator</li>
 +
<a href="https://webshop.fishersci.com/insight2_uk/getProduct.do?productCode=11626250&resultSetPosition=0"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Large Centrifuge</h3>
 +
<ul>
 +
<li>Eppendorf Centrifuge 5810 R</li>
 +
<a href="http://www.eppendorf.com/int/index.php?pb=01edea0fe7a0e358&action=products&contentid=1&catalognode=10019"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Nano Drop</h3>
 +
<ul>
 +
<li>Thermo Scientific NanoDrop® 2000 UV-Vis spectrophotometer combined with NanoDrop 2000 / 2000c Software</li>
 +
<a href="http://www.nanodrop.com/Productnd2000overview.aspx"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>PCR Machine</h3>
 +
<ul>
 +
<li>Applied Biosystems Veriti 96Well Thermal Cycler</li>
 +
<a href="https://www.thermofisher.com/ch/en/home/life-science/pcr/thermal-cyclers-realtime-instruments/thermal-cyclers/veriti-thermal-cycler.html"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Plate Reader</h3>
 +
<ul>
 +
<li>Tecan Infinite M200 Pro™ combined with Tecan iControl TM software</li>
 +
<a href="http://www.tecan.com/platform/apps/product/index.asp?MenuID=1812&ID=1916&Menu=1&Item=21.2.10.1"><p>for further information (platereader) click here</p></a>
 +
<a href="http://www.tecan.ch/platform/apps/product/index.asp?MenuID=1817&ID=1924&Menu=1&Item=21.7.8"><p>for further information (software) click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Shaker</h3>
 +
<ul>
 +
<li>Kuhner Climo-Shaker ISF1-X</li>
 +
<a href="http://www.kuhner.com/de/produkte/schuettelmaschinen/inkubatoren/isf1-x.html"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
 +
<div class="info">
 +
 +
<h3>Small Centrifuge</h3>
 +
<ul>
 +
<li>Eppendorf Centrifuge 5424 R</li>
 +
<a href="http://www.labdepotinc.com/Product_Details~id~450~pid~59833.aspx"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>UV- and Dark Hood</h3>
 +
<ul>
 +
<li>Biostep Dark-Hood DH-50™ combined with Argus-X1™ software and Biostep argusX1™ basic licence</li>
 +
<a href="http://www.biostep.de/i18n_en_DE/products/Bio_Imaging_components_1776/Tripod__dark_hoods_and_accessories_1863/Dark_hoods_1865/Dark_Hood_DH_40_with_control_panel_and_display_6/index.html"><p>for further information (hood) click here</p></a>
 +
<a href="https://www.biostep.de/products/Bio_Imaging_software_1843/Acquisition_and_control_software_1844/argus_X1___for_cameras_and_scanners_1845/biostep_argusX1__basic_licence_1/index.html"><p>for further information (software) click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
<div class="info">
 +
 +
<h3>Vortex</h3>
 +
<ul>
 +
<li>Scientific Industries VortexGenie®2</li>
 +
<a href="http://www.scientificindustries.com/genie2.html"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
<div class="info">
 +
 +
<h3>LI-COR Odyssey®</h3>
 +
<ul>
 +
<li>LI-COR Odyssey® CLx infrared Western Blot Scanner</li>
 +
<a href="http://www.licor.com/bio/products/imaging_systems/odyssey/"><p>for further information click here</p></a>
 +
</ul>
 +
<a class="expander" href="#" onclick="expand(this);return false;"><img src="https://static.igem.org/mediawiki/2015/1/1f/Blank_square.png"></a>
 +
</div>
 +
 +
 +
  
  
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{{:Template:ETH_Zurich/footer}}

Latest revision as of 23:53, 18 September 2015

"What I cannot create I do not understand."
- Richard Feynmann

Materials

Bacterial strains and Mammalian Cell Lines

Escherichia coli TOP10

The bacterial strain we used as a base for all our transformations was TOP10 (Genotype: F- mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 nupG recA1 araD139 Δ(ara-leu)7697 galE15 galK16 rpsL(StrR) endA1 λ-). These cells were provided by the Panke Group at ETH Zurich.

Jurkat Cells

Immortalized human T lymphocyte cell line. We used this cell line as a model line for cancer cells with elevated lactate production and TRAIL susceptibility (papers). A big thank you to the Tay Group at ETH Zurich who kindly offered us an aliquot of their cells.

HL60

Cell line derived from acute myeloid leukemia. We used this cell line as a model line for cancer cells with elevated lactate production and TRAIL susceptibility (papers). Many thanks to the Neri Group at ETH Zurich who kindly offered us an aliquot of their cells.

SK-BR-3

HER2-positive breast cancer cell line. We used this cell line to validate the chimeric receptors of the Stockholm iGEM team. We are very grateful to Michal Stanczak from the University Hospital of Basel for offering us a sample of this cell line.

3T3

Murine fibroblast cell line. We used this cell line as a model line for healthy cells which are restistant to TRAIL and produce normal levels of lactate. Many thanks to the Tay Group at ETH Zurich who kindly offered us an aliquot of their cells.

3T3 P65 -/-

Murine fluorescent fibroblast cell line.

  • H2B-GFP: nuclear staining
  • P65- dsRed: cytoskeleton
We used this cell line to study the co-culture of the bacteria and the chip. Many thanks to the Tay Group at ETH Zurich who kindly offered us an aliquot of their cells.

Buffers and media

Annexin V binding buffer

  • 10 mM HEPES
  • 140 mM NaCl
  • 2.5 mM
  • 250 mM CaCl2
  • pH 7.4
  • Filter the solution through a 0.2 µm filter.

Jurkat medium

  • 2 mL MEM 1x
  • 20 mL FBS 100%
  • 2 mL GlutaMax 100x
  • 0.4 mL Penicillin (10k units/mL) /Streptomyocin (10k µg/mL )
  • 200 mL RPMI

Terrific Broth

  • 2.32 g/L KH2PO4
  • 12.54 g/L K2HPO4
  • 1.2% Tryptone
  • 2.4% Yeast Extract
  • 0.4% Gylcerol
  • Filter the solution through a 0.2 µm filter.

SOC medium

  • 20 g/L Tryptophane
  • 5 g/L Yeast extract
  • 0.5 g/L NaCl
  • 250 mM KCl
  • 1 M MgCl2
  • 50% (w/v) sterile glucose
  • Filter the solution through a 0.2 µm filter.

TFB1

  • 100 mM RbCl
  • 50 mM MnCl2
  • 30 mM potassium acetate
  • 10 mM CaCl2
  • 15% glycerol

TFB2

  • 100 mM MOPS
  • 50 mM RbCl
  • 75 mM CaCl2
  • 15% glycerol

3T3 medium

  • 500 mL D-MEM 1x
  • 50 mL FBS 100%
  • 5 mL GlutaMax 100x
  • 5 mL Penicillin (10k units/mL) /Streptomyocin (10k µg/mL )

Chemicals

LB-Broth

Agarose

Agar

Glycerol

2-log DNA ladder

Taq DNA Ligase

Cutsmart Buffer

EcoRI

BsaI-HF

DpnI

Q5 Master Mix

dNTP Mix

Taq DNA Polymerase

AnnexinV-Alexa

Green DNA/RNA Dye

Protein G coated Beads

Streptavidin Beads

Red Gel Stain

Anti-Human-AnnexinV

Gel Loading Dye

T4 DNA Ligase

PstI

DNA Ligase Buffer

Ethanol

DMEM

DMEM with Glutamine

RPMI

Accutase

SuperKillerTRAIL™

TRAIL

Kits

Plasmid Purification

  • Product: ZR Plasmid Miniprep ™-Classic
  • Company: Zymo Research
  • Lot: ZRC182892
  • Catalog No: D4016

Changes to the protocol included an extra step to dry the column before elution and the use of water for elution instead of elution buffer. For elution of large plasmids we used prewarmed water.

Gel Extraction

  • Product: ZymocleanTM Gel DNA Recovery Kit
  • Company: Zymo Research
  • Lot: ZRC183055
  • Catalog No: D4002

Changes to the protocol included an extra step to dry the column before elution and the use of water for elution instead of elution buffer. For elution of large plasmids we used prewarmed water.

Lactate Detection Kit

  • Product: L-Lactic Acid
  • Company: Megazyme
  • Lot:
  • Catalog No:

Changes to the protocol included an extra step to dry the column before elution and the use of water for elution instead of elution buffer. For elution of large plasmids we used prewarmed water.

The detection of lactate using this kit relies on two enzymatic reactions. In the first, L-lactate and NAD&sup+; were converted into pyruvate and NADH+H&sup+; by L-lactate dehydrogenase. In the coupled reaction, pyruvate is used with D-glutamate by the D-glutamate-pyruvate transaminase to have the equilibrium of the first reaction in favour of the pyruvate. The concentration of NADH is finally measured as the optical density at 340 nm (OD340).

Machines

Absorbance Measurments

Balance

Gel Electrophoresis

Emulsifier

Eppi-shaker/Heatingblock

FACS

Incubator

Large Centrifuge

Nano Drop

PCR Machine

Plate Reader

Shaker

Small Centrifuge

UV- and Dark Hood

Vortex

LI-COR Odyssey®

We would like to thank our sponsors