Solutions/Media

LB Medium (1 L)
a) Measure out ~900 mL ddH₂O.
b) Add 10 g tryptone, 5 g yeast extract, and 10 g NaCl. Stir until all solids dissolve.
c) Adjust pH to 7.0 with 1 M NaOH.
d) Bring to volume with ddH₂O.
e) Autoclave Liquid Cycle 20 minutes at 15 psi (1.05 kg/cm²).

LB-Agar Plates (1 L of media=Forty 100 mm petri dishes)
a) Measure out ~900 mL ddH₂O.
b) Add 10 g tryptone, 5 g yeast extract, 15 g Bacto Agar, and 10 g NaCl. Stir.
c) Adjust pH to 7.0 with 1 M NaOH.
d) Bring to volume with ddH₂O.

e) Autoclave Liquid Cycle 20 minutes at 15 psi (1.05 kg/cm²).
f) Remove liquid from autoclave. Mix gently on stir plate to distribute melted agar (CAREFUL: superheated liquids may boil over when swirled).
g) Allow solution to cool to 50-60ºC. Add antibiotics.
h) Pour media into plates.
i) Allow media to harden.
j) Invert plates. Mark type of plate with marker. Place back in sleeve. Store at 4ºC.


NZM, NZYM, NZCYM Medium (1 L)
a) Measure out ~900 mL ddH₂O.
b) Add 10 g NZ amine (casein hydrolysate enzymatic), 5 g NaCl, and 2 g MgSO₄•7H₂O.
c) For NZYM medium, add 5 g yeast extract in addition to solutes in b.
d) For NZCYM medium, add 5 g yeast extract and 1 g casamino acids in addition to solutes in b.
e) Stir until all solutes dissolved.
f) Adjust pH to 7.0 with 1 M NaOH.
g) Adjust the volume to 1 L with ddH₂O.
h) Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).

2 M MgCl₂ (100 mL)
a) Measure out 90 mL ddH₂O.
b) Dissolve 19 g MgCl₂.
c) Adjust volume to 100 mL.
d) Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).

1 M Glucose (100 mL)
a) Measure out 90 mL ddH₂O.
b) Dissolve 18 g glucose.
c) Adjust volume to 100 mL.
d) Sterilize through 0.22 µm filter.


SOB, SOC Medium (1 L)
a) Measure out ~900 mL ddH₂O.
b) Add 20 g tryptone, 5 g yeast extract, and 0.5 g NaCl. Stir until all solutes dissolve.
c) Add 10 mL 250 mM KCl (Dissolve 1.86 g KCl in 100 mL ddH₂O to make 250 mM KCl).
d) Adjust pH to 7.0 with 1.0 M NaOH. Adjust volume to 1 L. Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).
e) If making SOC medium, allow solution to cool to 60ºC. Add 20 mL sterile 1M glucose.
f) Prior to use, add 5 mL sterile 2 M MgCl₂ for both SOB and SOC medium.

0.17 M KH₂PO₄/0.72 M K₂HPO₄ (100 mL)
a) Measure out 90 mL ddH₂O.
b) Dissolve 2.31 g KH₂PO₄ and 12.54 g K₂HPO₄.
c) Adjust volume to 100 mL.
d) Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).

Terrific Broth (1 L)
a) Measure out ~900 mL ddH₂O.
b) Add 12 g tryptone, 24 g yeast extract, and 4 mL glycerol. Stir until all solutes dissolve.
c) Adjust volume to 1 L. Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).
d) Allow solution to cool to 60ºC. Add 100 mL sterile 0.17 M KH₂PO₄/0.72 M K₂HPO₄.

2X YT Medium (1 L)
a) Measure out ~900 mL ddH₂O.
b) Add 16 g tryptone, 10 g yeast extract, and 5 g NaCl. Stir until solutes dissolve.
c) Adjust pH to 7.0 with 1 M NaOH. Adjust volume to 1 L.
d) Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).

Reference: Molecular Cloning, 3^rd Ed., Sambrook and Russell

PYE Media for growth of Caulobacter strains (1 L)
a)Measure out 950 mL ddH₂O.
b)Add 0.4 g beef extract, 0.2 g sodium acetate, 0.2 yeast extract, and 0.4 g tryptone.  Stir until solutes dissolve.
c)Adjust volume to 1 L.
d)Autoclave on liquid cycle 20 minutes at 15 psi (1.05 kg/cm²).