Team:Evry/Notebook/Protocols/Transformation

From 2014.igem.org

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<ul>
<ul>
-
  <li>Place electroporation tanks in ice for 10min</li><br>
+
  <li>Place electroporation tanks in ice for 10min</li>
-
<br>
+
  <li>Take samples of plasmids (25ng-50ng/µL)    /!\ Keep them in ice /!\ </li>
-
  <li>Take samples of plasmids (25ng-50ng/µL)    /!\ Keep them in ice /!\ </li><br>
+
  <li>Take sample of competent cells  /!\ Keep them in ice /!\ </li>
-
<br>
+
  <li>Place 1µL of plasmid in the sample of competent cells</li>  
-
  <li>Take sample of competent cells  /!\ Keep them in ice /!\ </li><br>
+
  <li>Transfer the full volume obtained in the electroporation tank</li>  
-
<br>
+
  <li>Place in the electroporator and pulse at 2000V </li>
-
  <li>Place 1µL of plasmid in the sample of competent cells</li> <br>
+
  <b>NB:</b>  The optimal pulse length is between 5 and 6ms.</li>
-
<br>
+
  <li>Add 1mL of MB 1X in the 30 seconds following the transformation</li>
-
  <li>Transfer the full volume obtained in the electroporation tank</li> <br>
+
  <li>Incubate between 2h and 3h at 30°C with shaking</li>
-
<br>
+
  <li>Centrifuge to concentrate all cells in the pellet</li>
-
  <li>Place in the electroporator and pulse at 2000V </li><br>
+
  <li>Discard the supernatant</li>
-
  <b>NB:</b>  The optimal pulse length is between 5 and 6ms.</li> <br>
+
  <li>Sowed the pellet on selective plates of MB 1X </li>
-
<br>
+
-
  <li>Add 1mL of MB 1X in the 30 seconds following the transformation</li> <br>
+
-
<br>
+
-
  <li>Incubate between 2h and 3h at 30°C with shaking</li> <br>
+
-
<br>
+
-
  <li>Centrifuge to concentrate all cells in the pellet</li> <br>
+
-
<br>
+
-
  <li>Discard the supernatant</li><br>
+
-
<br>
+
-
  <li>Sowed the pellet on selective plates of MB 1X </li><br>
+
<br>
<br>
</ul>
</ul>

Latest revision as of 00:48, 18 October 2014

Transformation of Pseudovibrio

  • Place electroporation tanks in ice for 10min
  • Take samples of plasmids (25ng-50ng/µL) /!\ Keep them in ice /!\
  • Take sample of competent cells /!\ Keep them in ice /!\
  • Place 1µL of plasmid in the sample of competent cells
  • Transfer the full volume obtained in the electroporation tank
  • Place in the electroporator and pulse at 2000V
  • NB: The optimal pulse length is between 5 and 6ms.
  • Add 1mL of MB 1X in the 30 seconds following the transformation
  • Incubate between 2h and 3h at 30°C with shaking
  • Centrifuge to concentrate all cells in the pellet
  • Discard the supernatant
  • Sowed the pellet on selective plates of MB 1X