Media & Antibiotics

LB

• Add:
  • 10 g/L NaCl
  • 10 g/L Bacto-Tryptone
  • 0.5 g/L Bacto-Yeast Extract
  • ddH2O

to a sterile pyrex bottle

• autoclave
• (add antibiotic when it reaches ~45°C)
• store at +4°C

LB Agar

• Add:
  • 10 g/L NaCl
  • 10 g/L Bacto-Tryptone
  • 0.5 g/L Bacto-Yeast Extract
  • 15 g/L Bacto-Agar
  • ddH2O

to a sterile 1L flask

• autoclave
• (add antibiotic when it reaches ~45°C, shake gently to avoid bubbles)
• pour into Petri plates
• let them polymerize for ~2-3h
• invert plates and wrap them with aluminium foil and store at +4°C

SOB

• Add:
  • 5 g/L Bacto-Yeast Extract
  • 20 g/L Bacto-Tryptone
  • 10mM NaCl
  • 2.5mM KCl
  • 10mM MgSO4
  • 10mM MgCl2

to a sterile pyrex bottle

• (optional: check that pH is ~6.8, otherwise adjust with NaOH)
• autoclave
• (add antibiotic when it reaches ~45°C)
• store at +4°C

SOC

• SOB+20mM (3.6 g/L) of glucose (add filter-sterilized (0.2um) glucose to autoclaved SOB).

M9 supplemented with glycerol (M9gly)

For 1L of medium, add:

• 716 ml of autoclaved (and cooled to Tamb) ddH2O
• 200 ul of autoclaved or filtered (0.2um) CaCl2 0.5 M
• 200 ml of autoclaved M9 salts 5x (dissolve 56.4 g in 1 liter ddH2O = 5x stock)
• 34 ml of filtered (0.2um) thiamine hydrochloride MW=337.27g/mol (340 mg in 34 ml)
• 20 ml of autoclaved MgSO4 0.1 M
• 20 ml of 10% autoclaved casamino acids (dissolve 50 g in 500 ml = 10% stock)
• 10 ml of autoclaved 40% glycerol as carbon source
• mix all the solutions in sterility (each solution must be completely dissolved!)
• (add antibiotic)
• store at +4°C, protected from light

NOTE:

• M9 salts 5x
• 10% casamino acids

can be stored at +4°C

• MgSO4 0.1 M
• CaCl2 0.5 M
• glycerol 40%

can be stored at room temperature or +4°C

• thiamine hydrochloride (LIGHT SENSITIVE) is one-shot and must be prepared each time