Team:EPF-Lausanne/Notebook

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Revision as of 12:25, 15 July 2008

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Notebook

You should make use of the calendar feature on the wiki and start a lab notebook. This may be looked at by the judges to see how your work progressed throughout the summer. It is a very useful organizational tool as well.

Find more information on how to use the calendar feature by going to the general calendar page.

Protocols

Reference : Most of these protocols come from http://openwetware.org

SOC medium : Super Original Catabolite Repressor

Bactotryptone 20g. Bacto-yeast extract 5g. NaCl 0.5g. 1M KCl 2.5ml ddH2O to 1000 ml Total Volume 1000ml Adjust pH to 7, with 10N NaOH. Autoclave to sterilize Add 20 ml of 1M glucose before use.

TE :

10xTE for 1 liter from stock solutions 10 ml 1M Tris-HCl pH 8.0 2 ml 0.5M EDTA pH 8.0 988 ml ddH2O

→ 10xTE is 10 mM Tris-HCl and 1 mM EDTA

   * For the Tris-HCl use Tris base and adjust to desired pH using HCl.

SOB : Super Original Broth

Used in growing bacteria for preparing chemically competent cells Ingredients

   * 0.5% (w/v) yeast extract
   * 2% (w/v) tryptone
   * 10 mM NaCl
   * 2.5 mM KCl
   * 20 mM MgSO4

Per liter:

   * 5 g yeast extract
   * 20 g tryptone
   * 0.584 g NaCl
   * 0.186 g KCl
   * 2.4 g MgSO4

!!! Adjust to pH 7.5 prior to use. This requires approximately 25 ml of 1M NaOH per liter.

CCMB80 buffer : For 1L

   * 10 mM KOAc pH 7.0 (10 ml of a 1M stock/L)
   * 80 mM CaCl2.2H2O (11.8 g/L)
   * 20 mM MnCl2.4H2O (4.0 g/L)
   * 10 mM MgCl2.6H2O (2.0 g/L)
   * 10% glycerol (100 ml/L)
   * adjust pH DOWN to 6.4 with 0.1N HCl if necessary
         o adjusting pH up will precipitate manganese dioxide from Mn containing solutions.
   * sterile filter and store at 4°C
   * slight dark precipitate appears not to affect its function

Antibiotics : For each antibiotic, solutions of 5 ml have been done and stored at -20°C (09.07.08)

Transformation

We used DH5-alpha cells from Bart's lab stock. We therefore followed the InVitrogen protocol.

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