http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&feed=atom&action=history
Team:Heidelberg/Notebook MaM - Revision history
2024-03-28T23:33:46Z
Revision history for this page on the wiki
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Chenchenzhu: /* Real-time RT-PCR */
2009-11-20T16:35:33Z
<p><span class="autocomment">Real-time RT-PCR</span></p>
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<td colspan='2' style="background-color: white; color:black;">Revision as of 16:35, 20 November 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Real-time RT-PCR ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Real-time RT-PCR ===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Cells were collected as a pellet. Cell number must excede 1*10<sup>5</sup>cells/tube and no more than 6*10<sup>6</sup>/tube due to column capacity. In order to obtain reliable results and reduce the variance during the extraction, reverse transcription and PCR, we did the measurement for each promoter with 12 replicates from an identical sample. Resuspend the pellet with 600µl RLT buffer containing ß-ME (1:<del class="diffchange diffchange-inline">1000</del>). QIAcube follows protocol "RNeasy Mini Animal Cell QIAshredder Version 3" using RNeasy Plant Mini Kit. mRNA from untransfected HeLa cells was extracted to be "Standard" to perform plate-to-plate correction.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Cells were collected as a pellet. Cell number must excede 1*10<sup>5</sup>cells/tube and no more than 6*10<sup>6</sup>/tube due to column capacity. In order to obtain reliable results and reduce the variance during the extraction, reverse transcription and PCR, we did the measurement for each promoter with 12 replicates from an identical sample. Resuspend the pellet with 600µl RLT buffer containing ß-ME (1:<ins class="diffchange diffchange-inline">100</ins>). QIAcube follows protocol "RNeasy Mini Animal Cell QIAshredder Version 3" using RNeasy Plant Mini Kit. mRNA from untransfected HeLa cells was extracted to be "Standard" to perform plate-to-plate correction.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After RNA is extracted, the extracts were put on ice immediately. 2 tubes of 5 µl aliquots were taken, One for measuring RNA concentration and one for gel electrophoresis. The RNA concentration was measured using NanoDrop under 230 nm. Each sample was measured 3 times. The gel electrophoresis was performed to confirm the RNA integrity because RNA degrade under many conditions. We used 1% agarose gel and 0.5x TAE buffer. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After RNA is extracted, the extracts were put on ice immediately. 2 tubes of 5 µl aliquots were taken, One for measuring RNA concentration and one for gel electrophoresis. The RNA concentration was measured using NanoDrop under 230 nm. Each sample was measured 3 times. The gel electrophoresis was performed to confirm the RNA integrity because RNA degrade under many conditions. We used 1% agarose gel and 0.5x TAE buffer. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Next step is the Real-time RT-PCR. We had different setup for different kits.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Next step is the Real-time RT-PCR. We had different setup for different kits.</div></td></tr>
</table>
Chenchenzhu
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=162368&oldid=prev
Aradema3: /* Induction */
2009-10-22T00:53:45Z
<p><span class="autocomment">Induction</span></p>
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<td colspan='2' style="background-color: white; color:black;">Revision as of 00:53, 22 October 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | VDR [[Team:Heidelberg/Notebook_MaM#References|[7]]] || activator || MCF-7 || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=D1530%7CSIGMA&N25=0&QS=ON&F=SPEC Vitamine D3] || 100 &mu;M || 1:1000/ 1:10000 || 100 nM/ 1 nM || DMEM+++ || 24 h, 48 h, 72 h, 96 h || TECAN read every 24 hours -> see induction: FACS ||</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | VDR [[Team:Heidelberg/Notebook_MaM#References|[7]]] || activator || MCF-7 || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=D1530%7CSIGMA&N25=0&QS=ON&F=SPEC Vitamine D3] || 100 &mu;M || 1:1000/ 1:10000 || 100 nM/ 1 nM || DMEM+++ || 24 h, 48 h, 72 h, 96 h || TECAN read every 24 hours -> see induction: FACS ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | c-Jun [[Team:Heidelberg/Notebook_MaM#References|[1]]] || activator || HeLa || [https://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=E4269%7CSIGMA&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&F=SPEC EGF] || 100&mu;g/mL || 1:1000 || 100ng/mL || <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline"> </del>||</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | c-Jun [[Team:Heidelberg/Notebook_MaM#References|[1]]] || activator || HeLa || [https://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=E4269%7CSIGMA&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&F=SPEC EGF] || 100&mu;g/mL || 1:1000 || 100ng/mL || <ins class="diffchange diffchange-inline">DMEM+++ </ins>|| <ins class="diffchange diffchange-inline">3h </ins>|| <ins class="diffchange diffchange-inline">after 3 h, EGF was removed and FACS measurement was carried out another 4 hours later </ins>||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|-</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|-</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|}</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|}</div></td></tr>
</table>
Aradema3
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=162199&oldid=prev
Aradema3: /* Induction */
2009-10-22T00:48:48Z
<p><span class="autocomment">Induction</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || MCF-7 || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=P4359%7CSIGMA&N25=0&QS=ON&F=SPEC Pifithrin-&alpha;] || 2 mM || 1:1000 || 2 &mu;M || minimal medium || 12 h || ||</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || MCF-7 || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=P4359%7CSIGMA&N25=0&QS=ON&F=SPEC Pifithrin-&alpha;] || 2 mM || 1:1000 || 2 &mu;M || minimal medium || 12 h || ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | AHR [[Team:Heidelberg/Notebook_MaM#References|[6]]] || activator || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=B1760%7CSIGMA&N5=Product%20No.%7CBRAND_KEY&F=SPEC Benzo-a-pyrene] || <del class="diffchange diffchange-inline"> </del>|| 1:10 000 || <del class="diffchange diffchange-inline"> </del>|| DMEM+++ || 18 h || ||</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | AHR [[Team:Heidelberg/Notebook_MaM#References|[6]]] || activator || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=B1760%7CSIGMA&N5=Product%20No.%7CBRAND_KEY&F=SPEC Benzo-a-pyrene] || <ins class="diffchange diffchange-inline">100 mM </ins>|| 1:10 000 || <ins class="diffchange diffchange-inline">10 &mu;M </ins>|| DMEM+++ || 18 h || ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || activator || MCF-7 || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?lang=en&N4=48599%7CSUPELCO&N5=SEARCH_CONCAT_PNO%6CBRAND_KEY&F=SPEC TCDD] || || 1:3000 || || DMEM++ || 48 h || ||</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || activator || MCF-7 || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?lang=en&N4=48599%7CSUPELCO&N5=SEARCH_CONCAT_PNO%6CBRAND_KEY&F=SPEC TCDD] || || 1:3000 || || DMEM++ || 48 h || ||</div></td></tr>
</table>
Aradema3
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=161981&oldid=prev
Aradema3: /* Induction */
2009-10-22T00:40:48Z
<p><span class="autocomment">Induction</span></p>
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<td colspan='2' style="background-color: white; color:black;">Revision as of 00:40, 22 October 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || MCF-7 || [https://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=07741%7CFLUKA&N25=0&QS=ON&F=SPEC Everolimus] || || 1:500 000 || || minimal medium || 24 h || without HIF-bag ||</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || MCF-7 || [https://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=07741%7CFLUKA&N25=0&QS=ON&F=SPEC Everolimus] || || 1:500 000 || || minimal medium || 24 h || without HIF-bag ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | SREBP [[Team:Heidelberg/Notebook_MaM#References|[2]]], [[Team:Heidelberg/Notebook_MaM#References|[8]]] [[Team:Heidelberg/Notebook_MaM#References|[9]]] || activator || HeLa || Lipid-depleted-Serum (LDS) || <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline">1:200 </del>|| <del class="diffchange diffchange-inline"> </del>|| DMEM++ || 2-3 days || put on cells after transfection and put back on after 3 h with HPCD ||</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | SREBP [[Team:Heidelberg/Notebook_MaM#References|[2]]], [[Team:Heidelberg/Notebook_MaM#References|[8]]] [[Team:Heidelberg/Notebook_MaM#References|[9]]] || activator || HeLa || Lipid-depleted-Serum (LDS) || <ins class="diffchange diffchange-inline">- </ins>|| <ins class="diffchange diffchange-inline">0,5% </ins>|| <ins class="diffchange diffchange-inline">- </ins>|| DMEM++ || 2-3 days || put on cells after transfection and put back on after 3 h with HPCD ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || activator || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=H107%7CSIGMA&N25=0&QS=ON&F=SPEC HPCD] || <del class="diffchange diffchange-inline"> </del>|| 1:100 || <del class="diffchange diffchange-inline"> </del>|| DMEM++ with LDS|| 3 h || add after cells were 2-3 days in DMEM++ +LDS ||</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || activator || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=H107%7CSIGMA&N25=0&QS=ON&F=SPEC HPCD] || <ins class="diffchange diffchange-inline">500 mg/ml </ins>|| 1:100 || <ins class="diffchange diffchange-inline">500 &mu;g/ml </ins>|| DMEM++ with LDS|| 3 h || add after cells were 2-3 days in DMEM++ +LDS ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=C8667%7CSIGMA&N25=0&QS=ON&F=SPEC Cholesterol] || <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline"> </del>|| DMEM+++ || 2-3 days || ||</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=C8667%7CSIGMA&N25=0&QS=ON&F=SPEC Cholesterol] || <ins class="diffchange diffchange-inline">10 mg/ml </ins>|| <ins class="diffchange diffchange-inline">1:1000 </ins>|| <ins class="diffchange diffchange-inline">10 &mu;g/ml </ins>|| DMEM+++ || 2-3 days || ||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=L7914%7CSIGMA&N25=0&QS=ON&F=SPEC LDL] || <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline"> </del>|| <del class="diffchange diffchange-inline"> </del>|| DMEM+++ || 2-3 days || ||</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | || inhibitor || HeLa || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?D7=0&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&N4=L7914%7CSIGMA&N25=0&QS=ON&F=SPEC LDL] || <ins class="diffchange diffchange-inline">0,5 mg/ml </ins>|| <ins class="diffchange diffchange-inline">1:50 </ins>|| <ins class="diffchange diffchange-inline">10 &mu;g/ml </ins>|| DMEM+++ || 2-3 days || <ins class="diffchange diffchange-inline">got LDL from Jessica Schilde -> self-made in Runz' lab </ins>||</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>|- style="background-color:#9BDDFF;" </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | NF&kappa;B[[Team:Heidelberg/Notebook_MaM#References|[1]]] [[Team:Heidelberg/Notebook_MaM#References|[10]]] || activator || U2-OS || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=T0157%7CSIGMA&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&F=SPEC&lang=null%3E TNF&alpha;] || 2,5 mM || 1:1000 || 2,5 &mu;M || DMEM++ || 10 h || || </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | NF&kappa;B[[Team:Heidelberg/Notebook_MaM#References|[1]]] [[Team:Heidelberg/Notebook_MaM#References|[10]]] || activator || U2-OS || [http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=T0157%7CSIGMA&N5=SEARCH_CONCAT_PNO%7CBRAND_KEY&F=SPEC&lang=null%3E TNF&alpha;] || 2,5 mM || 1:1000 || 2,5 &mu;M || DMEM++ || 10 h || || </div></td></tr>
</table>
Aradema3
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=158889&oldid=prev
Hannahuckelmann: /* Material */
2009-10-21T23:05:34Z
<p><span class="autocomment">Material</span></p>
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<td colspan='2' style="background-color: white; color:black;">Revision as of 23:05, 21 October 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>__NOTOC__</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>__NOTOC__</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>=<del class="diffchange diffchange-inline">Material</del>=</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>=<ins class="diffchange diffchange-inline">Materials & Methods</ins>=</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Kits==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Kits==</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{| class="wikitable centered" style="margin: 1em auto 1em auto"</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{| class="wikitable centered" style="margin: 1em auto 1em auto"</div></td></tr>
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Hannahuckelmann
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=151686&oldid=prev
MichaelBartoschek: /* Methods */
2009-10-21T19:05:20Z
<p><span class="autocomment">Methods</span></p>
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<td colspan='2' style="background-color: white; color:black;">Revision as of 19:05, 21 October 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Plasmid DNA and DNA fragments were extracted using the Gel Extraction Kit from Qiagen following the manufacture instructions. To check the purity and amount of extracted DNA an aliquot was analysed using a NanoDrop.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Plasmid DNA and DNA fragments were extracted using the Gel Extraction Kit from Qiagen following the manufacture instructions. To check the purity and amount of extracted DNA an aliquot was analysed using a NanoDrop.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Cell Culture==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Cell Culture==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== RA-PCR ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== RA-PCR ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>0.8µL Stop 3 (O.58)<br></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>0.8µL Stop 3 (O.58)<br></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Measurement and Screening ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Measurement and Screening ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>****60°C, 1 min</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>****60°C, 1 min</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In each plate, the 5 housekeeping genes were crossed with 12 replicates as well as Standard. eGFP was mixed with 12 replicates and RNase-free water as negative control. After the program finished, data was collected from OneStep software 2.0 associated to the StepOnePlus system. With a certain threshold (0.05), a series of Ct values were determined, then analyzed with MatLab.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In each plate, the 5 housekeeping genes were crossed with 12 replicates as well as Standard. eGFP was mixed with 12 replicates and RNase-free water as negative control. After the program finished, data was collected from OneStep software 2.0 associated to the StepOnePlus system. With a certain threshold (0.05), a series of Ct values were determined, then analyzed with MatLab.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== References ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== References ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[10] Treede I., Braun A., Jeliaskova P., Giese T., Füllekrug J., Griffiths G., Stremmel W. & Ehehalt R. TNF-α-induced up-regulation of pro-inflammatory cytokines is reduced by phosphatidylcholine in intestinal epithelial cells. ''BMC Gastroenterology'' 9:53 (2009)</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[10] Treede I., Braun A., Jeliaskova P., Giese T., Füllekrug J., Griffiths G., Stremmel W. & Ehehalt R. TNF-α-induced up-regulation of pro-inflammatory cytokines is reduced by phosphatidylcholine in intestinal epithelial cells. ''BMC Gastroenterology'' 9:53 (2009)</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">[[Team:Heidelberg/Notebook_MaM#Methods|[Back]]]</ins></div></td></tr>
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</table>
MichaelBartoschek
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=151643&oldid=prev
MichaelBartoschek: /* Material */
2009-10-21T19:03:17Z
<p><span class="autocomment">Material</span></p>
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<td colspan='2' style="background-color: white; color:black;">← Older revision</td>
<td colspan='2' style="background-color: white; color:black;">Revision as of 19:03, 21 October 2009</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Marker==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Marker==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Restriction Enzymes==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Restriction Enzymes==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Bacteria Growth Media==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Bacteria Growth Media==</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Team:Heidelberg/Notebook_MaM#<del class="diffchange diffchange-inline">Notebook_MaM</del>|[Back]]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Team:Heidelberg/Notebook_MaM#<ins class="diffchange diffchange-inline">Material</ins>|[Back]]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Eucaryotic cell lines==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Eucaryotic cell lines==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*''' [[Team:Heidelberg/Eukaryopedia#HeLa| HeLa]]'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*''' [[Team:Heidelberg/Eukaryopedia#HeLa| HeLa]]'''</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*''' [[Team:Heidelberg/Eukaryopedia#U2-OS| U2-OS]]'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*''' [[Team:Heidelberg/Eukaryopedia#U2-OS| U2-OS]]'''</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Cell Culture Media==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Cell Culture Media==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* 1 % Pen/Strep</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* 1 % Pen/Strep</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* Krebs-Henseleit-Buffer (Sigma-Aldrich [http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/1/k3753pis.Par.0001.File.tmp/k3753pis.pdf])<br></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* Krebs-Henseleit-Buffer (Sigma-Aldrich [http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/1/k3753pis.Par.0001.File.tmp/k3753pis.pdf])<br></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Oligos ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Oligos ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Oligos for LAM-PCR===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Oligos for LAM-PCR===</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Oligos for real-time RT-PCR===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Oligos for real-time RT-PCR===</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Plasmids ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Plasmids ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | P58 || intron in submission plasmid || || || Ampicillin</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | P58 || intron in submission plasmid || || || Ampicillin</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Instruments ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Instruments ==</div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Team:Heidelberg/Notebook_MaM#Material|[Back]]]</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Lab material ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Lab material ==</div></td></tr>
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MichaelBartoschek
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=151564&oldid=prev
MichaelBartoschek at 18:59, 21 October 2009
2009-10-21T18:59:59Z
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Team:Heidelberg/Notebook_MaM#<del class="diffchange diffchange-inline">Notebook_MaM</del>|[Back]]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Team:Heidelberg/Notebook_MaM#<ins class="diffchange diffchange-inline">Material</ins>|[Back]]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Enzymes==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Enzymes==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Bacteria==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Bacteria==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | 6/16-Well plates || Nunclon<sup>TM</sup> Surface || nunc<sup>TM</sup></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>| style="font-weight:bold;" | 6/16-Well plates || Nunclon<sup>TM</sup> Surface || nunc<sup>TM</sup></div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Team:Heidelberg/Notebook_MaM#Material|[Back]]]</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>= Methods =</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>= Methods =</div></td></tr>
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MichaelBartoschek
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=151505&oldid=prev
MichaelBartoschek at 18:57, 21 October 2009
2009-10-21T18:57:46Z
<p></p>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Team:Heidelberg/Notebook_MaM#Notebook_MaM|[Back]]]</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Eucaryotic cell lines==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Eucaryotic cell lines==</div></td></tr>
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MichaelBartoschek
http://2009.igem.org/wiki/index.php?title=Team:Heidelberg/Notebook_MaM&diff=151473&oldid=prev
MichaelBartoschek at 18:55, 21 October 2009
2009-10-21T18:55:51Z
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Enzymes==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Enzymes==</div></td></tr>
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MichaelBartoschek