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DSMZ

DSMZ 119 . METHANOBACTERIUM MEDIUM

Official DSMZ medium: This medium was manually curated by experts from the DSMZ.

Culture medium recipe

Vol.
×
Medium for Methanoculleus bourgensis DSM 6216

* Modification: Increase amount of Na-acetate to 3.00 g/l.

Growth conditions: anaerobic 37 °C

Main sol. 119 #

ml
Equipment needed: Hungate tubes Centrifuge Autoclave Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
KH2PO4 0.50 g 0.499 3.663
MgSO4 x 7 H2O 0.40 g 0.399 1.618
NaCl 0.40 g 0.399 6.824
NH4Cl 0.40 g 0.399 7.456
CaCl2 x 2 H2O 0.05 g 0.05 0.339
Trace element solution SL-10 1.00 ml - -
Yeast extract
(OXOID) 		1.00 g 0.997 -
Na-acetate* 3.00 g - -
Na-formate 2.00 g 1.994 29.321
FeSO4 x 7 H2O solution (0.1% w/v) 2.00 ml - -
Sludge fluid 50.00 ml - -
Fatty acid mixture 20.00 ml - -
Sodium resazurin
(0.1% w/v) 		0.50 ml 5.0e-4 0.002
NaHCO3 4.00 g 3.988 47.473
L-Cysteine HCl x H2O 0.50 g 0.499 2.838
Na2S x 9 H2O 0.50 g 0.499 2.076
Distilled water 930.00 ml - -
1 Dissolve ingredients except bicarbonate, cysteine and sulfide. Sparge medium with 80% H2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Add and dissolve bicarbonate, adjust pH to 6.5 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: 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.

Sludge fluid #

1000
Solution occurs in 7 other media (first defined in Medium 119).
Compound Amount Unit Conc. [g/L] Conc. [mM]
Yeast extract 4 g 4 -
Sludge 1000 ml - -
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 8-12°C in the dark.

Fatty acid mixture #

1000
Solution occurs in 6 other media (first defined in Medium 119).
Compound Amount Unit Conc. [g/L] Conc. [mM]
Isobutyric acid 23 ml - -
DL-2-Methylbutyric acid 27 ml - -
Valeric acid 27 ml - -
Isovaleric acid 27 ml - -
Distilled water 896 ml - -
1 Adjust pH to 7.5 with concentrated NaOH.

Trace element solution SL-10 #

1000
Solution occurs in 267 other media (first defined in Medium 320).
Compound Amount Unit Conc. [g/L] Conc. [mM]
HCl
(25%) 		10.0 ml 2.5 68.568
FeCl2 x 4 H2O 1.5 g 1.5 7.545
ZnCl2 70.0 mg 0.07 0.514
MnCl2 x 4 H2O 100.0 mg 0.1 0.505
H3BO3 6.0 mg 0.006 0.097
CoCl2 x 6 H2O 190.0 mg 0.19 1.463
CuCl2 x 2 H2O 2.0 mg 0.002 0.012
NiCl2 x 6 H2O 24.0 mg 0.024 0.185
Na2MoO4 x 2 H2O 36.0 mg 0.036 0.149
Distilled water 990.0 ml - -
1 First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml.

FeSO4 x 7 H2O solution (0.1% w/v) #

1000
Solution occurs in 13 other media (first defined in Medium 119).
Compound Amount Unit Conc. [g/L] Conc. [mM]
FeSO4 x 7 H2O 1 g 1 3.597
H2SO4
(0.1 N) 		1000 ml - -
1 The ferrous sulfate solution is not stable and should be freshly prepared.

Metadata

Last modified: 15.09.22
Source: DSMZ
Taxonomic range:  Archaea
Taxonomic range:  Bacteria
Medium type: Complex medium
Final pH: 6.8 - 7.0
Equipment needed: Autoclave, Centrifuge, Gassing station, Hungate tubes
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Gas composition

  • 80 % H2
  • 20 % CO2

Detailed instructions for the cultivation of anaerobes including important warnings can be found in the cultivation instructions .

Associated strains

All strains for this medium: 80
Strains with modifications: 22
Methanoculleus bourgensis DSM 6216

  Na-acetate
  3.00 g Na-acetate

Molecular composition
* modified beta

Compound Conc. [g/L] Conc. [mM]
NaHCO3 3.988 47.473
Na-acetate 2.991 36.461
Na-formate 1.994 29.321
KH2PO4 0.499 3.663
L-Cysteine HCl x H2O 0.499 2.838
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Cultivation metadata from 80 strains BacDive

pH 6 - 8 3 strains
pH 8 - 10 1 strain
pH 4 - 6 1 strain
phyla 4 84 strains
oxygen anaerobe 77 strains
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