Team:TU-Delft/Protocol 3

From 2013.igem.org

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<h4 align="left">Requirements:</h4>
<h4 align="left">Requirements:</h4>
<ol>
<ol>
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<li> Glycerol</li>
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<li>bacterial culture </li>
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<li> Pipettes</li>
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    <li>LB medium  </li>
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<li> 1.5 mL Tubes</li>
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    <li>80% glycerol  </li>
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<li>     Culture Sample</li>
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    <li>centrifuge  </li>
</ol>
</ol>
<br>
<br>
<h4 align="left">Prodecure:</h4>
<h4 align="left">Prodecure:</h4>
<ol>
<ol>
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<li> Label the tubes and add 180μL of glycerol. </li>
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<li> Pipette out 1000 μL of the culture. </li>
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    <li>Take 5 mL bacterial cells from the Erlenmeyer of a freshly grown culture and spin in a 15 mL tube for 10 minutes at 2,000 rpm (Eppendorf centrifuge). </li>
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<li> Add the culture to the tubes and store in -80°C</li>
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    <li>Decant the supernatant without disturbing the pellet. </li>
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    <li>Pipet on the pellet 0.5 mL 1% LB medium and 0.5 mL 80% glycerol and mix by vortexing and save in -80 °C freezer.  </li>
</ol>
</ol>
<br>
<br>

Revision as of 14:57, 3 October 2013



Protocols

Our project deals with E.coli cells which sense Auto-inducing peptides (AIPs) from the Staphylococcus aureus and starts producing Antimicrobial peptides in order to kill the Staphylococcus aureus. Different protocols used during the project are described below.




Making glycerol stocks

Requirements:

  1. bacterial culture
  2. LB medium
  3. 80% glycerol
  4. centrifuge

Prodecure:

  1. Take 5 mL bacterial cells from the Erlenmeyer of a freshly grown culture and spin in a 15 mL tube for 10 minutes at 2,000 rpm (Eppendorf centrifuge).
  2. Decant the supernatant without disturbing the pellet.
  3. Pipet on the pellet 0.5 mL 1% LB medium and 0.5 mL 80% glycerol and mix by vortexing and save in -80 °C freezer.

The glycerol stock can be used whenever required, by just adding 0.5 mL of stock into 5 mL of freshly prepared media.

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