Difference between revisions of "Team:Munich/purify1.html"

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<h4>T7 DNA extraction by phenol chloroform precipitation </h4>
+
<h4>T7 DNA Extraction by Phenol Chloroform Precipitation </h4>
 
<em>2018/05/17</em>
 
<em>2018/05/17</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
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       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno</td>
+
       <td>Keno Eilers</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td>T7 DNA Purification, phenol-chloroform precipitation</td>
+
       <td>
 +
<a href="https://static.igem.org/mediawiki/2018/7/7a/T--Munich--PhageT7DNAPurification.pdf" target="_blank">Phage T7 DNA Purification</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> still too much from E. coli
+
       <td> Still too much DNA from E. coli
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> DNA concentration: 55,5 ng/uL
+
       <td> DNA concentration: 55,5 ng/µL
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
 
   
 
   
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<figure class="figure">
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  <img src="SOURCE" class="figure-img img-fluid rounded" alt="error">
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    <figcaption class=" 0,3% Agarose gel, 2 log ladder ">OPTIONAL CAPTION</figcaption>
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    </figure> HIER FOTO
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</div>
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</html>
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<html>
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<div class="event-info">
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<h4>Cleanup of T7 DNA from 15.5.18 with NEB PCR Cleanup Kit </h4>
 
<h4>Cleanup of T7 DNA from 15.5.18 with NEB PCR Cleanup Kit </h4>
Line 55: Line 40:
 
   
 
   
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Nils</td>
+
       <td>Nils O'Brien</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td> NEB PCR-cleanup Kit, agarose gel </td>
+
       <td>
 +
<a href="https://www.neb.com/-/media/catalog/datacards-or-manuals/manualt1030.pdf" target="_blank">NEB PCR Cleanup Kit</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/b/be/T--Munich--WL1_Agarose-Gel_electrophoresis.pdf" target="_blank">Agarose Gel electrophoresis</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> warming of columns on 40 °C for a few minutes before elution </td>
+
       <td> Warming of columns on 40 °C for a few minutes before elution </td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> no results, not possible with genome larger than 25kb
+
       <td> No results, not possible with genome larger than 25kb
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
 
−
</html>
 
 
   
 
   
 
   
 
   
−
+
<h4>Second Try: Cleanup of T7 DNA from 15.5.18 with NEB PCR Cleanup Kit </h4>
−
<h4>second try: Cleanup of T7 DNA from 15.5.18 with NEB PCR Cleanup Kit </h4>
+
 
<em>2018/05/20</em>
 
<em>2018/05/20</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
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       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Nils</td>
+
       <td>Nils O'Brien</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td> NEB PCR-cleanup Kit, agarose gel </td>
+
       <td>
 +
<a href="https://www.neb.com/-/media/catalog/datacards-or-manuals/manualt1030.pdf" target="_blank">NEB PCR Cleanup Kit</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/b/be/T--Munich--WL1_Agarose-Gel_electrophoresis.pdf" target="_blank">Agarose Gel electrophoresis</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> warming of columns on 50 °C for a few minutes before elution </td>
+
       <td> Warming of columns on 50 °C for a few minutes before elution </td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> no results, did not work out for large fragments
+
       <td> No visible bands on 0,3 Agarose gel, did not work out for large fragments. <br>
 +
End of experiments with this method
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
−
+
 
   
 
   
 
   
 
   
−
<h4>T7 DNA clean up T7 DNA </h4>
+
<h4>T7 DNA Clean-Up T7 DNA </h4>
 
<em>2018/05/22</em>
 
<em>2018/05/22</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
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       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno, Kilian</td>
+
       <td>Keno Eilers</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td> phenol chloroform precipitation </td>
+
       <td>
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>
 +
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
        
−
      <td> no results à  testing PEG cutoff tomorrow
+
−
</td>
+
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
 
   
 
   
 +
 
   
 
   
−
+
<h4> Generating MS2 RNA Genome </h4>
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<html>
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<div class="event-info">
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<h4> Generating MS2 RNA genome </h4>
+
 
<em>2018/05/23</em>
 
<em>2018/05/23</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
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       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno</td>
+
       <td>Keno Eilers</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td>cDNA from PCR product, phenol chloroform precipitation </td>
+
       <td>  
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>,
 +
<a href="https://static.igem.org/mediawiki/2018/3/34/T--Munich--cDNASynthesis.pdf" target="_blank">
 +
cDNA Synthesis</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> 1 ul DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
+
       <td> 1 µl DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
 
       <td> RNA concentration: 34900 ng/µl
 
       <td> RNA concentration: 34900 ng/µl
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
 
   
 
   
−
+
 
−
+
<h4> Generating MS2 RNA Genome </h4>
−
<h4> Generating MS2 RNA genome </h4>
+
 
<em>2018/05/23</em>
 
<em>2018/05/23</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
Line 164: Line 150:
 
   
 
   
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno</td>
+
       <td>Keno Eilers</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td>cDNA from PCR product, phenol chloroform precipitation </td>
+
       <td>
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>,
 +
<a href="https://static.igem.org/mediawiki/2018/3/34/T--Munich--cDNASynthesis.pdf" target="_blank">
 +
cDNA Synthesis</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> 1 ul DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
+
       <td> 1 µl DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
 
       <td> RNA concentration: 34900 ng/µl
 
       <td> RNA concentration: 34900 ng/µl
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
<h4> Optimization of DNA Precipitation out of T7 Phage </h4>
−
+
<em>2018/05/23 - 2018/05/25</em>
−
<h4> Generating MS2 RNA genome </h4>
+
−
<em>2018/05/23</em>
+
 
<table class="table table-borderless">
 
<table class="table table-borderless">
 
     <tr>
 
     <tr>
 
   
 
   
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno</td>
+
       <td>Keno Eilers, Sophie Kurzbach, Nils O'Brien</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td>cDNA from PCR product, phenol chloroform precipitation </td>
+
       <td>
 +
<a href="https://static.igem.org/mediawiki/2018/7/7a/T--Munich--PhageT7DNAPurification.pdf" target="_blank">Phage T7 DNA Purification</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/b/be/T--Munich--WL1_Agarose-Gel_electrophoresis.pdf" target="_blank">Agarose Gelelectrophoresis</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> 1 ul DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
+
       <td> Step 11 of "T7 DNA purification" protocol: Optimization of PEG (63%) precipitation
 +
Incubation for 42 minutes, then proceeded with centrifugation
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> RNA concentration: 34900 ng/µl
+
       <td> 150 µl PEG: 302 ng/µl, 200 µl PEG: 164 ng/µl, 250 µl PEG: 278 ng/µl, 300 µl PEG: 113 ng/µl, 350 µl PEG: 279 ng/µl, 400 µl PEG: 216 ng/µl, 450 µl PEG: 530 ng/µl,  500 µl PEG: 422 ng/µl
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
 
   
 
   
−
<h4> Optimization of DNA precipitation out of T7 Phage </h4>
+
<h4>T7 DNA Isolation with Different Infection Times </h4>
−
<em>2018/05/23 - 2018/05/25</em>
+
<em>2018/05/29</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
 
     <tr>
 
     <tr>
 
   
 
   
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno, Sophie K., Nils</td>
+
       <td>Sophie Kurzbach</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td>T7 DNA purification, agarose gel, </td>
+
       <td>  
 +
<a href="https://static.igem.org/mediawiki/2018/7/7a/T--Munich--PhageT7DNAPurification.pdf" target="_blank">Phage T7 DNA Purification</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> Step 11 of "T7 DNA purification" protocol: Optimization of PEG (63%) precipitation
+
       <td> Tested transfection times: 12 min (before supernatant), 22 min (after supernatant), 32 min (cell lysis)
−
Incubation for 42 minutes, then proceeded with centrifugation
+
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> 150 µl PEG: 302 ng/µl, 200 µl PEG: 164 ng/µl, 250 µl PEG: 278 ng/µl, 300 µl PEG: 113 ng/µl, 350 µl PEG: 279 ng/µl, 400 µl PEG: 216 ng/µl, 450 µl PEG: 530 ng/µl, 500 µl PEG: 422 ng/µl
+
       <td> 12 min: 209 ng/µL, 22 min: 628 ng/µL, 32 min:  334 ng/µL
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
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<html>
+
−
<div class="event-info">
+
 
   
 
   
 
   
 
   
−
<h4>T7 DNA Isolation with different infection times </h4>
+
<h4> T7 DNA Isolation 10 min and 32 min after Super Transfection + RNAse </h4>
−
<em>2018/05/29</em>
+
<em>2018/06/05 -2018/06/06</em>
 
<table class="table table-borderless">
 
<table class="table table-borderless">
 
     <tr>
 
     <tr>
 
   
 
   
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Sophie K.</td>
+
       <td>Sophie Kurzbach</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td> T7 DNA purification, phenol chloroform precipitant </td>
+
       <td>  
 +
<a href="https://static.igem.org/mediawiki/2018/7/7a/T--Munich--PhageT7DNAPurification.pdf" target="_blank">Phage T7 DNA Purification</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>,
 +
PEG percipitant
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> tested transfection times: 12 min (before supernatant), 22 min (after supernatant), 32 min (cell lysis)
+
       <td> Better results than DNA without RNAse treatment, still too much E. coli DNA
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> 12 min: 209 ng/uL, 22 min:  628 ng/uL, 32 min:  334 ng/uL
+
       <td> DNA concentration of 10 min: 633 ng/µL
 +
DNA concentration of 32 min:  200 ng/µL
 
</td>
 
</td>
−
    </tr>
+
    </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
 
 
   
 
   
−
+
<h4>T7 DNA Extraction by Phenol Chloroform Precipitation </h4>
−
+
<em>2018/06/16-2018/06/18</em>
−
<html>
+
−
<div class="event-info">
+
−
+
−
+
−
<h4> T7 DNA Isolation 10 min and 32 min after super transfection + RNAse </h4>
+
−
<em>2018/06/05 -2018/06/06</em>
+
 
<table class="table table-borderless">
 
<table class="table table-borderless">
 
     <tr>
 
     <tr>
 
   
 
   
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Sophie K.</td>
+
       <td>Keno Eilers</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td> T7 DNA purification, PEG precipitant, phenol chloroform precipitant </td>
+
       <td>  
 +
<a href="https://static.igem.org/mediawiki/2018/7/7a/T--Munich--PhageT7DNAPurification.pdf" target="_blank">Phage T7 DNA Purification</a>,  
 +
<a href="https://static.igem.org/mediawiki/2018/0/0f/T--Munich--PhenolChloroformPrecipitation.pdf" target="_blank">
 +
Phenol Chloroform precipitation</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> better results than DNA without RNAse treatment, still too much E. coli DNA
+
       <td> No notes
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> DNA concentration of 10 min: 633 ng/uL
+
       <td> Good DNA: 10,37 nM
−
DNA concentration of 32 min:  200 ng/uL
+
 
</td>
 
</td>
−
 
−
<figure class="figure">
 
−
  <img src="SOURCE" class="figure-img img-fluid rounded" alt="error">
 
−
    <figcaption class=" 0,3% Agarose gel, 2 log ladder, after 10 min infection ">OPTIONAL CAPTION</figcaption>
 
−
    </figure> HIER FOTO
 
−
 
−
<figure class="figure">
 
−
  <img src="SOURCE" class="figure-img img-fluid rounded" alt="error">
 
−
    <figcaption class=" 0,3% Agarose gel, 2 log ladder, after 10 min infection (upper picture) and 32 min infection ">OPTIONAL CAPTION</figcaption>
 
−
    </figure> HIER FOTO
 
−
 
 
     </tr>
 
     </tr>
 
</table>
 
</table>
−
</div>
+
 
−
</html>
+
<h4> DNA Purification of T7 DNA with Qiagen Genomic Tips 100/G</h4>
−
+
<em>2018/06/22</em>
−
+
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<html>
+
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<div class="event-info">
+
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+
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+
−
<h4>T7 DNA extraction by phenol chloroform precipitation </h4>
+
−
<em>2018/06/16-2018/06/18</em>
+
 
<table class="table table-borderless">
 
<table class="table table-borderless">
 
     <tr>
 
     <tr>
 +
      <td>Participants:</td>
 +
      <td>Brigit Tunaj</td>
 +
    </tr>
 +
    <tr>
 +
      <td>Protocol:</td>
 +
      <td>
 +
<a href="http://algimed.by/download/EN-QIAGEN-Genomic-DNA-Handbook.pdf" target="_blank">Qiagen Genomic Tip Kit</a>
 +
</td>
 +
    </tr>
 +
    <tr>
 +
      <td>Notes:</td>
 +
      <td> Preparation of buffers as descripted on their homepage
 +
</td>
 +
    </tr>
 +
<tr>
 +
      <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
 +
      <td> Did not work out
 +
</td>
 +
    </tr>
 +
</table>
 
   
 
   
 +
<h4> DNA Purification of T7 DNA with Qiagen Genomic Tips 100/G</h4>
 +
<em>2018/06/25</em>
 +
<table class="table table-borderless">
 +
    <tr>
 
       <td>Participants:</td>
 
       <td>Participants:</td>
−
       <td>Keno</td>
+
       <td>Brigit Tunaj</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Protocol:</td>
 
       <td>Protocol:</td>
−
       <td> T7 DNA purification, phenol chloroform precipitant </td>
+
       <td>  
 +
<a href="http://algimed.by/download/EN-QIAGEN-Genomic-DNA-Handbook.pdf" target="_blank">Qiagen Genomic Tip Kit</a>
 +
</td>
 
     </tr>
 
     </tr>
 
     <tr>
 
     <tr>
 
       <td>Notes:</td>
 
       <td>Notes:</td>
−
       <td> -
+
       <td> Increased DNA concentration 50 ug
 
</td>
 
</td>
 
     </tr>
 
     </tr>
 
<tr>
 
<tr>
−
       <td>Results:</td>
+
       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
−
       <td> good DNA: 10,37 nM
+
       <td> Did not work out. <br>
 +
Method not suitable for phage DNA.
 
</td>
 
</td>
 
     </tr>
 
     </tr>

Latest revision as of 02:22, 18 October 2018

T7 DNA Extraction by Phenol Chloroform Precipitation

2018/05/17
Participants: Keno Eilers
Protocol: Phage T7 DNA Purification, Phenol Chloroform precipitation
Notes: Still too much DNA from E. coli
Results: DNA concentration: 55,5 ng/µL

Cleanup of T7 DNA from 15.5.18 with NEB PCR Cleanup Kit

2018/05/19
Participants: Nils O'Brien
Protocol: NEB PCR Cleanup Kit, Agarose Gel electrophoresis
Notes: Warming of columns on 40 °C for a few minutes before elution
Results: No results, not possible with genome larger than 25kb

Second Try: Cleanup of T7 DNA from 15.5.18 with NEB PCR Cleanup Kit

2018/05/20
Participants: Nils O'Brien
Protocol: NEB PCR Cleanup Kit, Agarose Gel electrophoresis
Notes: Warming of columns on 50 °C for a few minutes before elution
Results: No visible bands on 0,3 Agarose gel, did not work out for large fragments.
End of experiments with this method

T7 DNA Clean-Up T7 DNA

2018/05/22
Participants: Keno Eilers
Protocol: Phenol Chloroform precipitation

Generating MS2 RNA Genome

2018/05/23
Participants: Keno Eilers
Protocol: Phenol Chloroform precipitation, cDNA Synthesis
Notes: 1 µl DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
Results: RNA concentration: 34900 ng/µl

Generating MS2 RNA Genome

2018/05/23
Participants: Keno Eilers
Protocol: Phenol Chloroform precipitation, cDNA Synthesis
Notes: 1 µl DNAse I (NEB, 2.000 U/mL) added, incubate for 10 minutes at 37 °C
Results: RNA concentration: 34900 ng/µl

Optimization of DNA Precipitation out of T7 Phage

2018/05/23 - 2018/05/25
Participants: Keno Eilers, Sophie Kurzbach, Nils O'Brien
Protocol: Phage T7 DNA Purification, Agarose Gelelectrophoresis
Notes: Step 11 of "T7 DNA purification" protocol: Optimization of PEG (63%) precipitation Incubation for 42 minutes, then proceeded with centrifugation
Results: 150 µl PEG: 302 ng/µl, 200 µl PEG: 164 ng/µl, 250 µl PEG: 278 ng/µl, 300 µl PEG: 113 ng/µl, 350 µl PEG: 279 ng/µl, 400 µl PEG: 216 ng/µl, 450 µl PEG: 530 ng/µl, 500 µl PEG: 422 ng/µl

T7 DNA Isolation with Different Infection Times

2018/05/29
Participants: Sophie Kurzbach
Protocol: Phage T7 DNA Purification, Phenol Chloroform precipitation
Notes: Tested transfection times: 12 min (before supernatant), 22 min (after supernatant), 32 min (cell lysis)
Results: 12 min: 209 ng/µL, 22 min: 628 ng/µL, 32 min: 334 ng/µL

T7 DNA Isolation 10 min and 32 min after Super Transfection + RNAse

2018/06/05 -2018/06/06
Participants: Sophie Kurzbach
Protocol: Phage T7 DNA Purification, Phenol Chloroform precipitation, PEG percipitant
Notes: Better results than DNA without RNAse treatment, still too much E. coli DNA
Results: DNA concentration of 10 min: 633 ng/µL DNA concentration of 32 min: 200 ng/µL

T7 DNA Extraction by Phenol Chloroform Precipitation

2018/06/16-2018/06/18
Participants: Keno Eilers
Protocol: Phage T7 DNA Purification, Phenol Chloroform precipitation
Notes: No notes
Results: Good DNA: 10,37 nM

DNA Purification of T7 DNA with Qiagen Genomic Tips 100/G

2018/06/22
Participants: Brigit Tunaj
Protocol: Qiagen Genomic Tip Kit
Notes: Preparation of buffers as descripted on their homepage
Results: Did not work out

DNA Purification of T7 DNA with Qiagen Genomic Tips 100/G

2018/06/25
Participants: Brigit Tunaj
Protocol: Qiagen Genomic Tip Kit
Notes: Increased DNA concentration 50 ug
Results: Did not work out.
Method not suitable for phage DNA.