Team:TU-Delft/Protocol 3

From 2013.igem.org

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<li>bacterial culture </li>
<li>bacterial culture </li>
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    <li>LB medium </li>
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  <li>80% glycerol  </li>
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    <li>80% glycerol </li>
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    <li>centrifuge </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> The bacterial cells are grown overnight.  </li>
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     <li>Decant the supernatant without disturbing the pellet.  </li>
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     <li> In a glass tube take 250 μL of 80 % Glycerol solution.  </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>
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     <li> Pipet 1mL of the cell culture into 0.25 mL 80% glycerol and mix by vortexing and save in -80 °C freezer.  </li>
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Revision as of 15:00, 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. 80% glycerol

Prodecure:

  1. The bacterial cells are grown overnight.
  2. In a glass tube take 250 μL of 80 % Glycerol solution.
  3. Pipet 1mL of the cell culture into 0.25 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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