1315 .
METHANOGEN MS-I MEDIUM
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Equipment needed:
Anaerobic workbench
Autoclave
Filter
Final pH:
7.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
NaCl | 0.450 | g | ||
|
|
NaHCO3 | 0.200 | g | ||
|
|
MgSO4 x 7 H2O | 0.090 | g | ||
|
|
KH2PO4 | 0.225 | g | ||
|
|
K2HPO4 | 0.225 | g | ||
|
|
(NH4)2SO4 | 0.225 | g | ||
|
|
CaCl2 x 2 H2O | 0.060 | g | ||
|
|
(NH4)2Ni(SO4)2 x 6 H2O | 2.000 | mg | ||
|
|
FeSO4 x 7 H2O | 2.000 | mg | ||
|
|
Wolfe's mineral elixir | 0.100 | ml | ||
|
|
Resazurin | 1.000 | mg | ||
|
|
Distilled water | 1000.000 | ml | ||
| 1 Mix components thoroughly, except NaHCO3, and adjust pH to 7.0. Bring to a boil for a seconds, cool down under a stream of N2-CO2 (80:20, v/v) and add NaHCO3 to the medium. Dispense the medium into culture vessels (e.g., 20 ml in 120 ml serum bottles) under a H2-CO2 (80:20, v/v) gas mixture, seal with butyl rubber stoppers and autoclave. Aseptically and anaerobically add per liter the following solutions (autoclaved or filter-sterilized, and stored under N2): | |||||
|
|
Trace vitamins
(*) |
10.000 | ml | ||
|
|
Sodium acetate
(1.0 M) |
10.000 | ml | ||
|
|
Na2S x 9 H2O
(5%) |
10.000 | ml | ||
| 2 Pressurize inoculated bottles to 100 KPa H2-CO2 (80:20, v/v). | |||||
Wolfe's mineral elixir #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
MgSO4 x 7 H2O | 30.00 | g | ||
|
|
MnSO4 x n H2O | 5.00 | g | ||
|
|
NaCl | 10.00 | g | ||
|
|
FeSO4 x 7 H2O | 1.00 | g | ||
|
|
CoCl2 x 6 H2O | 1.80 | g | ||
|
|
CaCl2 x 2 H2O | 1.00 | g | ||
|
|
ZnSO4 x 7 H2O | 1.80 | g | ||
|
|
CuSO4 x 5 H2O | 0.10 | g | ||
|
|
AlK(SO4)2 x 12 H2O | 0.18 | g | ||
|
|
H3BO3 | 0.10 | g | ||
|
|
Na2MoO4 x 2 H2O | 0.10 | g | ||
|
|
(NH4)2Ni(SO4)2 x 6 H2O | 2.80 | g | ||
|
|
Na2WO4 x 2 H2O | 0.10 | g | ||
|
|
Na2SeO4 | 0.10 | g | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 First adjust pH to 1.0 with diluted H2SO4, then dissolve the salts. | |||||
Trace vitamins #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
Biotin | 2.0 | mg |
|
|
Folic acid | 2.0 | mg |
|
|
Pyridoxine hydrochloride | 10.0 | mg |
|
|
Thiamine HCl | 5.0 | mg |
|
|
Riboflavin | 5.0 | mg |
|
|
Nicotinic acid | 5.0 | mg |
|
|
Calcium pantothenate | 5.0 | mg |
|
|
Vitamin B12 | 0.1 | mg |
|
|
p-Aminobenzoic acid | 5.0 | mg |
|
|
Lipoic acid | 5.0 | mg |
|
|
Distilled water | 1000.0 | ml |