Medium data

Search for medium no. = [265].
Unless otherwise stated, sterilize media by autoclaving at 121°C for 15 min.

265   METHANOGENIUM MEDIUM

KCl  0.355   g
MgCl2·6H2O  4.0     g
MgSO4·7H2O  3.45    g
NH4Cl  0.25    g
CaCl2·2H2O  0.14    g
KH2PO4  0.14    g
NaCl  18.0     g
Wolfe's mineral solution (see below)  10.0     ml
Trace vitamins (see Medium No. 197)  10.0     ml
Fe(NH4)2(SO4)2·6H2O  2.0     mg
NaHCO3  5.0     g
Sodium acetate  1.0     g
Yeast extract (BD-Difco)  2.0     g
Trypticase peptone (BD-BBL)  2.0     g
Resazurin  1.0     mg
Distilled water  980.0     ml
Mix components thoroughly, except NaHCO3, and adjust pH to 6.5. Bring to a boil for 5-10 sec, then, cool down under a stream of H2-CO2 (80:20, v/v) and add NaHCO3 to the medium. Dispense the medium into suitable culture vessels (e.g., 5 ml medium in Hungate tubes) under the same gas mixture and seal with butyl rubber stoppers. After autoclaving, stand the medium overnight. Aseptically and anaerobically add per liter the following solutions (autoclaved and stored under a N2 atmosphere):
5% L-Cysteine·HCl·H2O solution  10.0     ml
5% Na2S·9H2O solution  10.0     ml
Pressurize the inoculated bottles to 200 kPa H2:CO2 (80:20, v/v).
Wolfe's mineral solution:
Trace minerals (see Medium No. 151)  1.0     L
NiCl2·6H2O  0.02    g
Na2SeO3  0.001   g
Na2WO4·2H2O  0.01    g