IGEM:PennState/2006/Growthmedia

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Contents

Growth Media

LB

Media (per liter)

  • 10 g Bactotryptone
  • 5 g Yeast Extract
  • 10 g NaCl
  • pH to 7.0 with NaOH/HCl
  • Autoclave

Plates

  • Add 15 g Bacto-Agar per liter media
  • Autoclave for 25 minutes
    • If antibiotics necessary, add only once media has cooled to ~45ºC, or is comfortable to the touch
  • Pour in sterile environment (~15ml per plate)
  • Cool in such a way as to avoid accumulation of condensation, while maintaining sterility
  • Seal with parafilm, place in bag, and store at 4ºC

LBM

Media (per liter)

  • 10 g Bactotryptone
  • 5 g Yeast Extract
  • 5 g NaCl
  • 0.4066 g MgCl2.6H2O (or 2mL 1M MgCl2)
  • pH to 7.0 with NaOH/HCl
  • Autoclave

Plates

  • Add 15 g Bacto-Agar per liter media
  • Autoclave for 25 minutes
    • If antibiotics necessary, add only once media has cooled to ~45ºC, or is comfortable to the touch
  • Pour in sterile environment (~15ml per plate)
  • Cool in such a way as to avoid accumulation of condensation, while maintaining sterility
  • Seal with parafilm, place in bag, and store at 4ºC

Top Agar (per liter)

  • 500 mL LBM
  • 500 mL DI H2O
  • 8 g Bacto-Agar
  • Autoclave

SOB

Media (per liter)

  • 20 g bactotryptone
  • 5 g bacto-yeast extract
  • 0.5 g NaCl
  • 0.19 g KCl
  • pH to 7.0 with NaOH/HCl
  • Autoclave for 25 minutes
  • Add filter-sterilized MgCl2 and MgSO4 to yield final concentration of 10 mM each

Final Concentration:

  • 0.5% Yeast Extract
  • 2% Tryptone
  • 10mM NaCl
  • 2.5 mM KCl
  • 10 mM MgCl2
  • 10 mM MgSO4

SOC

Media

  • Add filter-sterilized glucose to SOB for 20 mM final concentration of glucose

Minimal Media

M9 Salts (per liter)

  • 1. Make M9 salts
  • 2. To make M9 Salts aliquot 800ml H2O and add
    • 64g Na2HPO4-7H2O
    • 15g KH2PO4
    • 2.5g NaCl
    • 5.0g NH4Cl
    • Stir until dissolved
    • Adjust to 1000ml with distilled H2O
    • Sterilize by autoclaving

Media (per liter)

  • 3. Measure ~700ml of distilled H2O (sterile)
  • 4. Add 200ml of M9 salts
  • 5. Add 2ml of 1M MgSO4 (sterile)
  • 6. Add 20 ml of 20% glucose (or other carbon source)
  • 7. Add 100ul of 1M CaCl2 (sterile)
  • 8. Adjust to 1000ml with distilled H2O
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