Team:Freiburg Bioware/NoteBook/Labjournal

From 2010.igem.org

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<div class="box_labjournal_195">
<div class="box_labjournal_195">
<a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/March">
<a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/March">
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<h2 style="margin-left: 5px;">April</h2>
<h2 style="margin-left: 5px;">April</h2>
<p class="boxes_labjournal_day">Labday 2- 5</p>
<p class="boxes_labjournal_day">Labday 2- 5</p>
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<img src="https://static.igem.org/mediawiki/2010/f/fc/Freiburg_10_labjournal_april_v1.JPG" class="image_labjournal" title="Scanning for iGEM restriction sites:<br/>
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<img src="https://static.igem.org/mediawiki/2010/f/fc/Freiburg_10_labjournal_april_v1.JPG" class="image_labjournal" title="Scanning for iGEM restriction sites:  
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• Vector plasmid <br/>
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Vector plasmid; Thymidine kinase ;Cytosindeaminase" />
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• Thymidine kinase<br/>
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• Cytosindeaminase" />
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<p class="boxes_labjournal_text">...ordering AAV-2 Helper-free System...</p>
<p class="boxes_labjournal_text">...ordering AAV-2 Helper-free System...</p>
</a>
</a>
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<p class="boxes_labjournal_day">Labday 18- 45</p>
<p class="boxes_labjournal_day">Labday 18- 45</p>
<img src="https://static.igem.org/mediawiki/2010/3/37/Freiburg_10_labjournal_june_v1.jpg" class="image_labjournal" title="
<img src="https://static.igem.org/mediawiki/2010/3/37/Freiburg_10_labjournal_june_v1.jpg" class="image_labjournal" title="
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We conducted our first site-directed mutagenesis. <br/>
+
We conducted our first site-directed mutagenesis. First cell culture steps: Splitting and seeding of AAV 293 and HT1080 cells; Calcium phosphate transfection; Transduction with virus particles containing YFP encoding sequence; Microscopy: Fluorescent transduced HT1080 cells. We planned to determine infectious virus titer via quantitative real-time PCR. For this purpose primers were designed, transduced HT1080 cells were harvested and prepared. Theoretical studies of the AAV structure: Impressions" />
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First cell culture steps:<br/>
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• Splitting and seeding of AAV 293 and HT1080 cells<br/>
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-
• Calcium phosphate transfection<br/>
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-
•      Transduction with virus particles containing YFP encoding sequence<br/>
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-
• Microscopy: Fluorescent transduced HT1080 cells<br/>
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-
<br/>
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We planned to determine infectious virus titer via quantitative real-time PCR. For this purpose primers were designed, transduced HT1080 cells were harvested and prepared.<br/>
+
-
<br/>
+
-
Theoretical studies of the AAV structure: Impressions" />
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<p class="boxes_labjournal_text">...first transduced cells...</p>
<p class="boxes_labjournal_text">...first transduced cells...</p>
</a>
</a>
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<p class="boxes_labjournal_day">Labday 46- 75</p>
<p class="boxes_labjournal_day">Labday 46- 75</p>
<img src="https://static.igem.org/mediawiki/2010/8/8d/Freiburg_10_labjournal_july_v1.JPG" class="image_labjournal" title="We developed a plan for modifying the surface exposed loops of the virus capsid.
<img src="https://static.igem.org/mediawiki/2010/8/8d/Freiburg_10_labjournal_july_v1.JPG" class="image_labjournal" title="We developed a plan for modifying the surface exposed loops of the virus capsid.
-
For this purpose not only 5 iGEM restriction sites, but also the so called ViralBrick restriction sites located in the Rep/Cap sequence needed to be deleted: BamHI, SalI <br/>
+
For this purpose not only 5 iGEM restriction sites, but also the so called ViralBrick restriction sites located in the Rep/Cap sequence needed to be deleted: BamHI, SalI. BioBrick production of the inverted terminal repeats (ITRs) turned out to be difficult. Therefore the alternative “ITR fancy method” was developed. We successfully produced AAV2 left and right ITR fused to the RFC10 prefix. Further on we received first quantitative transduction results via flow cytometry. mGMK_TK30 was successfully cloned into the vector plasmid." />
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<br/>
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BioBrick production of the inverted terminal repeats (ITRs) turned out to be difficult. Therefore the alternative “ITR fancy method” was developed. We successfully produced AAV2 left and right ITR fused to the RFC10 prefix.<br/>
+
-
<br/>
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Further on we received first quantitative transduction results via flow cytometry.<br/>
+
-
<br/>
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mGMK_TK30 was successfully cloned into the vector plasmid." />
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<p class="boxes_labjournal_text">...organizing the lab...</p>
<p class="boxes_labjournal_text">...organizing the lab...</p>
</a>
</a>
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<h2 style="margin-left: 5px;">September II</h2>
<h2 style="margin-left: 5px;">September II</h2>
<p class="boxes_labjournal_day">Labday 124- 135</p>
<p class="boxes_labjournal_day">Labday 124- 135</p>
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<img src="https://static.igem.org/mediawiki/2010/3/35/Freiburg_10_BioBrick_icon.png" class="image_labjournal" />
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<img src="https://static.igem.org/mediawiki/2010/1/18/Freiburg_10_labjournal_october2_v1.JPG" class="image_labjournal" />
<p class="boxes_labjournal_text">...21x mutated RepCap is working...</p>
<p class="boxes_labjournal_text">...21x mutated RepCap is working...</p>
</a>
</a>
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<h2 style="margin-left: 5px;">October II</h2>
<h2 style="margin-left: 5px;">October II</h2>
<p class="boxes_labjournal_day">Labday 150- 166</p>
<p class="boxes_labjournal_day">Labday 150- 166</p>
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<img src="https://static.igem.org/mediawiki/2010/3/35/Freiburg_10_BioBrick_icon.png" class="image_labjournal" />
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<img src="https://static.igem.org/mediawiki/2010/2/22/Freiburg10_Oktober.png" class="image_labjournal" />
<p class="boxes_labjournal_text">...Capsid modification works...</p>
<p class="boxes_labjournal_text">...Capsid modification works...</p>
</a>
</a>
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<h2 style="margin-left: 5px;">November</h2>
<h2 style="margin-left: 5px;">November</h2>
<p class="boxes_labjournal_day">Labday 166- 170</p>
<p class="boxes_labjournal_day">Labday 166- 170</p>
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<img src="https://static.igem.org/mediawiki/2010/3/35/Freiburg_10_BioBrick_icon.png" class="image_labjournal" />
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<img src="https://static.igem.org/mediawiki/2010/9/93/Freiburg10_November.png" class="image_labjournal" />
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<p class="boxes_labjournal_text">...Jemboree!!!...</p>
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<p class="boxes_labjournal_text">...Jamboree!!!...</p>
</a>
</a>
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<br><h2>Get to know our virus</h2>
 
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<!---Insert Text in here---> Our project starts with first sequence analysis.
 
</div>
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<div class="lab_journal_img"><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/March" ><img src="https://static.igem.org/mediawiki/2010/2/21/Freiburg10_Labjournal-March.png" /></a></div></div>
 
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<div class="lab_journal_text">
 
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<br><h2>Checking iGEM compatibility </h2>
 
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<!---Insert Text in here--->Scanning for iGEM restriction sites:<br/>
 
-
• Vector plasmid <br/>
 
-
• Thymidine kinase<br/>
 
-
• Cytosindeaminase<br/>
 
-
 
-
</div>
 
-
<div class="lab_journal_img"><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/April" ><img src="https://static.igem.org/mediawiki/2010/6/6b/Freiburg10_Labjournal-April.png" /></a></div></div>
 
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<div class="lab_journal_box">
 
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<div class="lab_journal_text">
 
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<br><h2>First practical steps</h2>
 
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<!---Insert Text in here--->We took a closer look to the theoretical DNA- and amino acid sequence of the ITRs, questioned the role of the β-globin intron, had a closer look to the CMV promoter and performed our first cloning attempt.
 
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</div>
 
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<div class="lab_journal_img"><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/May" ><img src="https://static.igem.org/mediawiki/2010/6/61/Freiburg10_Labjournal-May.png" /></a></div></div>
 
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<div class="lab_journal_box">
 
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<div class="lab_journal_text">
 
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<br><h2>First fluorescing HT1080 cells</h2>
 
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<!---Insert Text in here--->We conducted our first site-directed mutagenesis. <br/>
 
-
<br/>
 
-
First cell culture steps:<br/>
 
-
• Splitting and seeding of AAV 293 and HT1080 cells<br/>
 
-
• Calcium phosphate transfection<br/>
 
-
•      Transduction with virus particles containing YFP encoding sequence<br/>
 
-
• Microscopy: Fluorescent transduced HT1080 cells<br/>
 
-
<br/>
 
-
We planned to determine infectious virus titer via quantitative real-time PCR. For this purpose primers were designed, transduced HT1080 cells were harvested and prepared.<br/>
 
-
<br/>
 
-
Theoretical studies of the AAV structure: Impressions.
 
-
 
-
</div>
 
-
<div class="lab_journal_img"><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/June" ><img src="https://static.igem.org/mediawiki/2010/4/48/Freiburg10_Labjournal-June.png" /></a></div></div>
 
-
 
-
<div class="lab_journal_box">
 
-
<div class="lab_journal_text">
 
-
<br><h2>Modifying Rep/Cap & Modularizing Vector plasmid</h2>
 
-
<!---Insert Text in here--->We developed a plan for modifying the surface exposed loops of the virus capsid.
 
-
For this purpose not only 5 iGEM restriction sites, but also the so called ViralBrick restriction sites located in the Rep/Cap sequence needed to be deleted: BamHI, SalI <br/>
 
-
<br/>
 
-
 
-
BioBrick production of the inverted terminal repeats (ITRs) turned out to be difficult. Therefore the alternative “ITR fancy method” was developed. We successfully produced AAV2 left and right ITR fused to the RFC10 prefix.<br/>
 
-
<br/>
 
-
Further on we received first quantitative transduction results via flow cytometry.<br/>
 
-
<br/>
 
-
mGMK_TK30 was successfully cloned into the vector plasmid.
 
-
 
-
</div>
 
-
<div class="lab_journal_img"><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/July" ><img src="https://static.igem.org/mediawiki/2010/d/dc/Freiburg10_Labjournal-July.png" /></a></div></div>
 
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<br><h2>Producing more BioBricks</h2>
 
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<!---Insert Text in here--->
 
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<div class="lab_journal_img"><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/August" ><img src="https://static.igem.org/mediawiki/2010/0/06/Freiburg10_Labjournal-AugustI.png" /></a></div></div>
 
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<br><h2>Headline</h2>
 
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https://2010.igem.org/Team:Freiburg_Bioware/NoteBook => Back to Notebook overview]
 
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<ul>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/March">March (labday 1)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/April">April (labday 2 - 5)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/May">May (labday 6 - 17)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/June">June (labday 18 - 45)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/July">July (labday 46 - 75)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/August">August part 1 (labday 76 - 92)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/August2">August part 2 (labday 93 - 106)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/September">September part 1 (labday 107 - 123)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/September2">September part 2 (labday 124 - 135)</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/October">October part 1 (labday 136 - 149 )</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/October2">October part 2 (labday 150 - 166 )</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/November">November  (labday 167 - 170 )</a></li>
 
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<li><a href="https://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal/Cellculture">Cellculture</a></li>
 
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{{:Team:Freiburg_Bioware/Footer}}
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Latest revision as of 03:17, 28 October 2010

March

Labday 1

...watching gene sequences...

April

Labday 2- 5

...ordering AAV-2 Helper-free System...

May

Labday 6- 17

...theoretical cloning...

June

Labday 18- 45

...first transduced cells...

July

Labday 46- 75

...organizing the lab...

August I

Labday 76- 92

...cloning parts...

August II

Labday 93- 106

...assembled vector plasmid is working...

September I

Labday 107- 123

...simply producing BioBricks...

September II

Labday 124- 135

...21x mutated RepCap is working...

October I

Labday 136- 149

...testing BioBricks by virus production...

October II

Labday 150- 166

...Capsid modification works...

November

Labday 166- 170

...Jamboree!!!...

Retrieved from "http://2010.igem.org/Team:Freiburg_Bioware/NoteBook/Labjournal"