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119. METHANOBACTERIUM MEDIUM

Culture medium recipe

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* Modified for Methanothermobacter tenebrarum DSM 23052
beta

Anaerobic

Solution occurs in 1 other media (first defined in medium 119 )
Equipment needed: Hungate tubes Autoclave
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.50 g 0.498 3.66
2 0.40 g 0.398 1.616
3 0.40 g 0.398 6.817
4 0.40 g 0.398 7.448
5 0.05 g 0.05 0.339
6
(0.1% w/v in 0.1 N H2SO4)
2.00 ml 0.002 0.007
7 Trace element solution SL-10 1.00 ml - -
8
(OXOID)
1.00 g 0.996 -
9 , might need strain adjustment 1.00 g 0.996 12.142
10 2.00 g 1.992 29.292
11 Sludge fluid 50.00 ml - -
12 Fatty acid mixture 20.00 ml - -
13
(0.1% w/v)
0.50 ml 5.0e-4 0.002
14 4.00 g 3.984 47.426
15 0.50 g 0.498 2.835
16 0.50 g 0.498 2.073
17 930.00 ml 924.634 51324.9
19 , might need strain adjustment 2.00 g/l 2 14.08
20 , might need strain adjustment 2.00 g/l 2 -
22 Vitamin solution , might need strain adjustment 10.00 ml/l - -
Instructions:
  1. Dissolve ingredients except bicarbonate, cysteine and sulfide. Sparge medium with 80% H2 and 20% CO2 gas mixture for 30 - 45 minto make it anoxic. Add and dissolve bicarbonate, adjust pH to 6.8 and dispense medium under 80% H2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials to 30% of their volume and autoclave. Add cysteine and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas. Prior to use check pH of complete medium and adjust to 6.8 - 7.0, if necessary.
  2. Note: The ferrous sulfate solution is not stable and should be freshly prepared. After growth has started and the culture is becoming turbid add sterile 80% H2 and 20% CO2 gas mixture to 0.5 - 1 bar overpressure.
Solution occurs in 8 other media (first defined in medium 119 )
Equipment needed: Autoclave Anaerobic workbench Centrifuge
Instructions:
  1. Add 0.4% yeast extract to sludge from an anaerobic digester, and after gassing with nitrogen gas for a few minutes incubate it at 37°C for 24 hours Then centrifuge the sludge at 13000 g and autoclave the resulting, clear supernatant in screw-capped vessels under nitrogen gas. The sludge fluid can be stored at room temperature in the dark.
Solution occurs in 10 other media (first defined in medium 119 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 23 ml - -
2 27 ml - -
3 27 ml 22.989 225.092
4 27 ml - -
5 1000 ml 904.173 50189.2
Instructions:
  1. Adjust pH to 7.0 - 7.3 with 2 N NaOH.
Solution occurs in 32 other media (first defined in medium 1310 )
Equipment needed: Centrifuge Filter Autoclave
Instructions:
  1. Rumen fluid from cow or sheep (obtained from fistulated animals or abattoir refuse) is filtered through muslin, autoclaved at 121°C for 15 minand then centrifuged at 27,000 g for 20 min The supernatant is made anoxic by sparging with 100% N2 gas for 15 min dispensed under same gas atmosphere into anoxic serum vials to 30% of volume and then stored frozen at - 20°C.
Solution occurs in 366 other media (first defined in medium 141 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 2.0 mg 0.002 0.008
2 2.0 mg 0.002 0.005
3 10.0 mg 0.01 0.049
4 5.0 mg 0.005 0.015
5 5.0 mg 0.005 0.013
6 5.0 mg 0.005 0.041
7 5.0 mg 0.005 0.01
8 0.1 mg 1.0e-4 6.3e-5
9 5.0 mg 0.005 0.036
10 5.0 mg 0.005 0.024
11 1000.0 ml 998.207 55408.8
Solution occurs in 239 other media (first defined in medium 320 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
2
(25%; 7.7 M)
10.0 ml 3.17 86.943
3 1.5 g 1.5 7.545
4 70.0 mg 0.07 0.514
5 100.0 mg 0.1 0.505
6 6.0 mg 0.006 0.097
7 190.0 mg 0.19 0.799
8 2.0 mg 0.002 0.012
9 24.0 mg 0.024 0.101
10 36.0 mg 0.036 0.149
11 990.0 ml 988.225 54854.8
Instructions:
  1. First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.0 ml.

Metadata

Media status: Official DSMZ medium
Source: DSMZ
Taxonomic range:  Archaea
Taxonomic range:  Bacteria
Medium type: Complex medium
Final pH: 6.9
Equipment needed: Anaerobic workbench, Autoclave, Centrifuge, Filter, Hungate tubes

Associated strains

All strains for this medium: 78
Strains with modifications: 78
Methanothermobacter tenebrarum DSM 23052

Gas composition

  • 20 % CO2
  • 80 % H2

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Distilled water 943.63 52379.3
NaHCO3 3.984 47.426
Sodium formate 1.992 29.292
Yeast extract 0.996 -
KH2PO4 0.498 3.66
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Cultivation metadata from 78 strains BacDive

halophily not 4 strains
halophily slightly 2 strains
pH 6 - 8 4 strains
pH 8 - 10 3 strains
pH 4 - 6 3 strains
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119a. METHANOBREVIBACTER MEDIUM 83 %
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