Team:Goettingen/Project/Materials

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Recipes

Media


Recipes for liquid and solid media for bacterial cultivation.
  1. LB-Medium
    2. 1% (w/v) bacto tryptone
    0.5% (w/v) bacto yeast extract
    1% (w/v) NaCl
    if required: ad 1% (w/v) agar

  2. M9 Swimming Agar
    3. 1.25% (v/v) glycerol
    20% (v/v) 5x M9 salt stock solution
    0.1% (v/v) of CaCl2 - 2H2O stock solution (20 mg/mL)
    0.1% (v/v) MgSO4 stock solution (0.12 g/mL)
    0.3 % (w/v) of agar (0.3 % [w/v])

  3. Tryptone swimming agar
    4. 1% (w/v) bacto tryptone
    0.5% (w/v) NaCl
    0.3% (w/v) agar


Antibiotics


Stock solutions and concentrations of antibiotics.


Supplements


Stock solutions and concentrations of supplements.


Buffer and Solution


Recipes for stocks of buffers and solutions.
  1. 5x M9 salts stock solution
    2. Stir until dissolved; sterilize by autoclaving at 121°C.
    Amount Substance
    64 g Na2HPO4 × 7 H2O
    15 g KH2PO4
    2.5 g NaCl
    5.0 g NH4Cl
    ad 1000 ml ddH2O


  2. CaCl2 Buffer for competent cells
    3. Stir until dissolved; sterilize by sterile filtration.
    Amount Substance Notes
    7.351 g CaCl2 × 7 H2O Alternative 0.1 M CaCl2.
    15% Glycerol
    ad 500 ml ddH2O


  3. 50x TAE for agarose gels
    4. Stir until dissolved. Working concentration is 1x (40 ml of 50x stock ad 2 l ddH2O).
    Amount Substance
    242 g Tris base
    57.1 ml Glacial acetic acid
    18.6 g EDTA
    ad 1000 ml ddH2O


E. coli

Strains


E. coli strain list - List of the working E. coli strains including genotypes, genetic markers, and alleles.


Storage


Storage of E. coli strains - How to keep E. coli strains.
  1. Glycerol Stock
    • 900 µl over night culture in LB-liquid medium
    • 900 µl 50% sterile glycerol
    Glycerol is extremely viscous. Hence, use cut off tips to pipette. Glycerol stocks were vortexed to ensure uniformly mixture and shock frozen in N2 or dry ice. Then, cultures were stored at -80°C. Do NOT forget to fill in the according GMO S1 construction sheet!


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