Difference between revisions of "Team:Pitt/Notebook"

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.notebook th {
 
.notebook th {
 
     border-bottom: 2px solid black;
 
     border-bottom: 2px solid black;
 +
width:50%;
 
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   <tr>   
 
   <tr>   
−
     <td>Crude lysate</td>
+
     <td>Crude lysate preparation from ERT7/DH5a</td>
 
     <td>Original S30 extract protocol, much too dilute due to incorrect weighing</td>
 
     <td>Original S30 extract protocol, much too dilute due to incorrect weighing</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> June 25, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Sequencing of Gibson'd protease constructs</td>
 +
    <td>5/6 had correct sequence, other one had significant deletion</td>
 +
  </tr>
 +
 +
  <tr> 
 +
    <td>ERT7 lysate tests on Tecan plate reader</td>
 +
    <td>No results because of high dilution</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> June 29, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Repetition of Pardee experiments</td>
 +
    <td>First success!</td>
 +
  </tr>
 +
 +
  <tr> 
 +
    <td>Freeze-dry constructs on paper</td>
 +
    <td>Test validity of Pardee results (Tatyana Yatsenko, Garrett Green)</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/><h4> June 30, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Grow O/N of NiCo21(DE3)</td>
 +
    <td>Competent cell prep (Konstantin Borisov)</td>
 +
  </tr>
 +
 +
  <tr> 
 +
    <td>Test freeze-dried constructs</td>
 +
    <td>No results on Tecan reader</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 1, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>NiCo21(DE3) competent cell prep</td>
 +
    <td>20 tubes of 250ul each (Konstantin Borisov)</td>
 +
  </tr>
 +
 +
  <tr> 
 +
    <td>Solution tests of Pardee toehold sensors</td>
 +
    <td>No results</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 6, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Restocked solutions, plates and media</td>
 +
    <td>Running low on supplies</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 7, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Transform plasmids for ERT7: wildtype T7, T7-YFP, T7 with sites</td>
 +
    <td>Controls for estrogen project, into DH5a</td>
 +
  </tr>
 +
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 9, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Created cell lysates from NiCo21(DE3), NiCo21(DE3) with IPTG induction, DH5a, DH5a with ERT7-YRP</td>
 +
    <td>Lots of controls</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 10, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Set up PCR for vector backbone</td>
 +
    <td>Discovered original vector backbone had been left out and degraded</td>
 +
  </tr>
 +
 +
  <tr> 
 +
    <td>Prepared glycerol stocks of protease constructs</td>
 +
    <td>In NiCo21(DE3) cells</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/><h4> July 11, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Gel electrophoresis on PCR</td>
 +
    <td>Gel was run in the wrong direction... lost all DNA</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 13, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Another PCR of vector backbone and Gel</td>
 +
    <td>Success! 3 lanes of vector backbone extracted.</td>
 +
  </tr>
 +
 +
  <tr> 
 +
    <td>Crude lysate preparation from ERT7/DH5a</td>
 +
    <td>Original S30 extract protocol, much too dilute due to incorrect weighing</td>
 +
  </tr>
 +
</table>
 +
<hr>
 +
<br/>
 +
<h4> July 16, 2015</h4>
 +
<table class="notebook">
 +
  <tr class="head">
 +
    <th>Protocol</th>
 +
    <th>Notes/results/purpose</th>
 +
  </tr>
 +
 
 +
  <tr> 
 +
    <td>Gibson of MMP9 fusion and protease reporter, transformation into DH5a</td>
 +
    <td>all transformations had a lot of colonies</td>
 
   </tr>
 
   </tr>
 
</table>
 
</table>

Revision as of 00:05, 19 September 2015

Notebook

Note: all experiments were done with all members present, unless otherwise noted

June 15, 2015

Protocol Notes/results/purpose
Transformation of plasmids from Pardee paper Into DH5a, all transformations successful
Grow O/N of DH5a Competent cell prep (Konstantin Borisov)

June 16, 2015

Protocol Notes/results/purpose
O/N of transformations Miniprep
DH5a competent cell prep 25 tubes of 200ul each, enough to last all summer! (Konstantin Borisov, help from Tatyana Yatsenko)

June 17, 2015

Protocol Notes/results/purpose
Transformation of plasmids from iGEM registry Into new DH5a, all transformations successful

June 18, 2015

Protocol Notes/results/purpose
Test PT7 lysate with Pardee paper plasmids No convenient method of visualization, so we started looking for a plate reader

June 22, 2015

Protocol Notes/results/purpose
PCR pSB1C3 with backbone fwd and rev primers For Gibson assembly, learning process
Gel of PCR products Visualization and purification of pSB1C3 backbone
Gel purificationObtained enough backbone for the rest of the summer!

June 23, 2015

Protocol Notes/results/purpose
Gibson assembly of protease parts from IDT, transformation into DH5a All transformations multiple colonies
Transformation of CMU ERT7 into DH5a Success, faint yellow from YFP seen under blue light

June 24, 2015

Protocol Notes/results/purpose
Miniprep and diagnostic gel of Gibson assembly All 3 had appropriate bands
Crude lysate preparation from ERT7/DH5a Original S30 extract protocol, much too dilute due to incorrect weighing

June 25, 2015

Protocol Notes/results/purpose
Sequencing of Gibson'd protease constructs 5/6 had correct sequence, other one had significant deletion
ERT7 lysate tests on Tecan plate reader No results because of high dilution

June 29, 2015

Protocol Notes/results/purpose
Repetition of Pardee experiments First success!
Freeze-dry constructs on paper Test validity of Pardee results (Tatyana Yatsenko, Garrett Green)

June 30, 2015

Protocol Notes/results/purpose
Grow O/N of NiCo21(DE3) Competent cell prep (Konstantin Borisov)
Test freeze-dried constructs No results on Tecan reader

July 1, 2015

Protocol Notes/results/purpose
NiCo21(DE3) competent cell prep 20 tubes of 250ul each (Konstantin Borisov)
Solution tests of Pardee toehold sensors No results

July 6, 2015

Protocol Notes/results/purpose
Restocked solutions, plates and media Running low on supplies

July 7, 2015

Protocol Notes/results/purpose
Transform plasmids for ERT7: wildtype T7, T7-YFP, T7 with sites Controls for estrogen project, into DH5a

July 9, 2015

Protocol Notes/results/purpose
Created cell lysates from NiCo21(DE3), NiCo21(DE3) with IPTG induction, DH5a, DH5a with ERT7-YRP Lots of controls

July 10, 2015

Protocol Notes/results/purpose
Set up PCR for vector backbone Discovered original vector backbone had been left out and degraded
Prepared glycerol stocks of protease constructs In NiCo21(DE3) cells

July 11, 2015

Protocol Notes/results/purpose
Gel electrophoresis on PCR Gel was run in the wrong direction... lost all DNA

July 13, 2015

Protocol Notes/results/purpose
Another PCR of vector backbone and Gel Success! 3 lanes of vector backbone extracted.
Crude lysate preparation from ERT7/DH5a Original S30 extract protocol, much too dilute due to incorrect weighing

July 16, 2015

Protocol Notes/results/purpose
Gibson of MMP9 fusion and protease reporter, transformation into DH5a all transformations had a lot of colonies