http://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&feed=atom&action=historyTeam:Tuebingen/Notebook - Revision history2024-03-28T22:12:39ZRevision history for this page on the wikiMediaWiki 1.16.0http://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=141104&oldid=prevJakobmatthes: /* Lab Notebook */2012-09-22T15:12:59Z<p><span class="autocomment">Lab Notebook</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookPreparations|Preparations]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookPreparations|Preparations]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookReports|Reports]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookReports|Reports]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookAppendix|Appendix]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookAppendix|Appendix]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookCalendar|Calendar]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>* [[Team:Tuebingen/NotebookCalendar|Calendar]]</div></td></tr>
</table>Jakobmattheshttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=141034&oldid=prevJakobmatthes: /* Lab Notebook */2012-09-22T15:08:34Z<p><span class="autocomment">Lab Notebook</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{{:Team:Tuebingen/Template/Tuebingen}}</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{{:Team:Tuebingen/Template/Tuebingen}}</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">=</del>= Lab Notebook <del class="diffchange diffchange-inline">=</del>=</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>= Lab Notebook =</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>''The following pages hold a detailed and technical account of the developement of our research project.''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>''The following pages hold a detailed and technical account of the developement of our research project.''</div></td></tr>
</table>Jakobmattheshttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=130037&oldid=prevJakobmatthes at 14:32, 20 September 20122012-09-20T14:32:49Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Lab Notebook ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Lab Notebook ==</div></td></tr>
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</table>Jakobmattheshttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=111298&oldid=prevJakobmatthes: /* Lab Notebook */2012-09-13T09:46:38Z<p><span class="autocomment">Lab Notebook</span></p>
<a href="http://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=111298&oldid=111245">Show changes</a>Jakobmattheshttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=111245&oldid=prevMiriW at 09:20, 13 September 20122012-09-13T09:20:59Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After determining our principal project idea we had to design our system prior to any work in the wet lab. Several steps were involved:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After determining our principal project idea we had to design our system prior to any work in the wet lab. Several steps were involved:</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Identification of plasmids ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Identification of plasmids ====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Due to several EcoRI and PstI restriction sites in the pRS plasmids (not located around the multiple cloning site), we can only use XbaI and SpeI for assembly.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Due to several EcoRI and PstI restriction sites in the pRS plasmids (not located around the multiple cloning site), we can only use XbaI and SpeI for assembly.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Identification of genes ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Identification of genes ====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Reporter:''' The targets of our signalling system regulate our reporter. We have access to the plasmid storage of our lab. Common reporters used in this yeast-based environment are firefly luciferase and beta-galactosidase. Both are available to us and have no legal issues concerning the publishing in the PartsRegistry.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Reporter:''' The targets of our signalling system regulate our reporter. We have access to the plasmid storage of our lab. Common reporters used in this yeast-based environment are firefly luciferase and beta-galactosidase. Both are available to us and have no legal issues concerning the publishing in the PartsRegistry.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Sequence analysis and primer design ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Sequence analysis and primer design ====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>The firefly luciferase has an unwanted XbaI site so we had to use NheI/SpeI restriction instead. The XbaI site could be removed by gene synthesis.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>The firefly luciferase has an unwanted XbaI site so we had to use NheI/SpeI restriction instead. The XbaI site could be removed by gene synthesis.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Weekly Reports ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Weekly Reports ===</div></td></tr>
</table>MiriWhttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=111241&oldid=prevMiriW at 09:20, 13 September 20122012-09-13T09:20:18Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>3. Preparing chemocompetent E. coli TOP 10 cells after Inoue protocol.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>3. Preparing chemocompetent E. coli TOP 10 cells after Inoue protocol.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 2 (7/16-7/22) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 2 (7/16-7/22) ====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>3. A new PCR of the parts 1-8 with Taq/Pfu-polymerase was performed applicating new yeast DNA. As an effect of the frequent freezing and defrosting the old yeast DNA was probably destroyed. Therefore some earlier PCRs did not work.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>3. A new PCR of the parts 1-8 with Taq/Pfu-polymerase was performed applicating new yeast DNA. As an effect of the frequent freezing and defrosting the old yeast DNA was probably destroyed. Therefore some earlier PCRs did not work.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 5 (8/06-8/12) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 5 (8/06-8/12) ====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>A Preparative gel for PCR products 3,4,5,6,7,8 (from PCR with Herculase) delivered new template for another PCR with Taq/Pfu Polymerase.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>A Preparative gel for PCR products 3,4,5,6,7,8 (from PCR with Herculase) delivered new template for another PCR with Taq/Pfu Polymerase.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 6 (8/13-8/19) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 6 (8/13-8/19) ====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>3. We received the synthesized receptor of Xenopus laevis. It was successfully transformed in E. coli TOP10 and purified with midi-prep.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>3. We received the synthesized receptor of Xenopus laevis. It was successfully transformed in E. coli TOP10 and purified with midi-prep.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 7 (8/20-8/26) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 7 (8/20-8/26) ====</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;">1. The PCR of the parts 1 and 2 with Herculase polymerase was executed.</ins></div></td></tr>
<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;">The PCR products were checked with an analytical gel afterwards. The PCR of part 1 failed again, so we decided to reject part 1 and continue working only with luciferase (part 2), because we only need one reporter gene.</ins></div></td></tr>
<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;"></ins></div></td></tr>
<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;">2. Since we ran out of luciferase DNA, we decided to transform remaining DNA of luciferase into E.coli TOP10. The transformation was successful.</ins></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;">3. The receptors (mPR Danio rerio, mPR Xenopus laevis) and mig1 were initially ligated into pRS vectors and transformed into E.coli (TOP10). But there were no colonies on the agar-plate.</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 8 (8/27-9/02) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 8 (8/27-9/02) ====</div></td></tr>
</table>MiriWhttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=111234&oldid=prevMiriW: /* Week 6 (8/13-8/19) */2012-09-13T09:17:04Z<p><span class="autocomment">Week 6 (8/13-8/19)</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 6 (8/13-8/19) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 6 (8/13-8/19) ====</div></td></tr>
<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;"></ins></div></td></tr>
<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;">1. The first successful ligation and transformation into pGEM vector of part 4 in E. Coli TOP10 was executed. A lot of colonies grew on the agar-plate. After a restriction digest with XbaI and SpeI and the control with a gel-electrophoresis the right band of 711bp was visible. The sequencing of the DNA confirmed that part 4 has the expected DNA-sequence.</ins></div></td></tr>
<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;">A midi-prep and restriction digest followed in order to prepare them for later ligation in pRS vectors.</ins></div></td></tr>
<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;"></ins></div></td></tr>
<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;">2. Ligation of part 3, 6, 7 and 8 in pGEM vector was performed. Reaction took place over night at 4 °C (39.2 Fahrenheit). The transformation of these parts was executed into E. coli TOP10.</ins></div></td></tr>
<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;">After growth over night, a mini plasmid preparation was performed. After a colony-PCR with parts 3,6,7,8 did not work, we had to get back to the restriction digest. Positive samples were prepared for sequencing. The Parts 3 and 8 were sequenced successfully. The ligation of parts 6 and 7 failed, so we decided to skip part 6, because we may use Psuc2 as promotor for luciferase. </ins></div></td></tr>
<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;"> </ins></div></td></tr>
<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;">3. We received the synthesized receptor of Xenopus laevis. It was successfully transformed in E. coli TOP10 and purified with midi-prep.</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 7 (8/20-8/26) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 7 (8/20-8/26) ====</div></td></tr>
</table>MiriWhttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=111232&oldid=prevMiriW: /* Week 5 (8/06-8/12) */2012-09-13T09:16:29Z<p><span class="autocomment">Week 5 (8/06-8/12)</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Therefore the plasmids (pRS313, 315, 316 and the parts Mig1, mPR D.R.) were purified again with a midi DNA purification kit. Now the restriction digest was executed completely.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Therefore the plasmids (pRS313, 315, 316 and the parts Mig1, mPR D.R.) were purified again with a midi DNA purification kit. Now the restriction digest was executed completely.</div></td></tr>
<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;">A mini plasmid preparation was performed afterwards to purify DNA in order to prepare the DNA for ligation.</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2. A new PCR of the parts 1-8 was executed using Herculase in order to obtain a higher amount of PCR-product. The polymerase Herculase was used due to its preciseness and productivity. Indeed the result of the PCR was better than with Pfu/Taq-polymerase.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2. A new PCR of the parts 1-8 was executed using Herculase in order to obtain a higher amount of PCR-product. The polymerase Herculase was used due to its preciseness and productivity. Indeed the result of the PCR was better than with Pfu/Taq-polymerase.</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">A Preparative gel for PCR products 3,4,5,6,7,8 (from PCR with Herculase) delivered new template for another PCR with Taq/Pfu Polymerase.</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 6 (8/13-8/19) ====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==== Week 6 (8/13-8/19) ====</div></td></tr>
</table>MiriWhttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=105882&oldid=prevLukZim at 14:44, 10 September 20122012-09-10T14:44:31Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{{:Team:Tuebingen/Template/Tuebingen}}</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{{:Team:Tuebingen/Template/Tuebingen}}</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">== Lab Notebook ==</del></div></td><td colspan="2"> </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;">== Lab Notebook ==</ins></div></td></tr>
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</table>LukZimhttp://2012.igem.org/wiki/index.php?title=Team:Tuebingen/Notebook&diff=105881&oldid=prevLukZim at 14:44, 10 September 20122012-09-10T14:44:25Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Lab Notebook ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Lab Notebook ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>__TOC__</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>__TOC__</div></td></tr>
</table>LukZim