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DSMZ

DSMZ 120b . METHANOMICROCOCCUS MEDIUM

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

Culture medium recipe

Vol.
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Medium for Methanimicrococcus hongohii DSM 114388

* Modification: Supplement medium after autoclaving with 0.5 g/l DTT (DL-dithiothreitol) added from an anoxic stock solution sterilized by filtration. Omit Na2S x 9 H2O and L-cysteine HCl x H2O.

Growth conditions: 1-2 days anaerobic 37 °C

Main sol. 120b #

ml
Equipment needed: Autoclave Hungate tubes Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
K2HPO4 0.35 g 0.342 1.962
KH2PO4 0.23 g 0.225 1.651
NH4Cl 0.50 g 0.488 9.128
MgSO4 x 7 H2O 0.50 g 0.488 1.981
CaCl2 x 2 H2O 0.25 g 0.244 1.661
NaCl 2.25 g 2.197 37.599
FeSO4 x 7 H2O solution (0.1% w/v) 2.00 ml - -
Trace element solution SL-10 1.00 ml - -
Yeast extract
(OXOID) 		2.00 g 1.953 -
Casitone
(BD BBL) 		2.00 g 1.953 -
Na-acetate 2.50 g 2.441 29.761
Sodium resazurin
(0.1% w/v) 		0.50 ml 4.9e-4 0.002
NaHCO3 2.50 g 2.441 29.062
2-Mercaptoethanesulfonate
(coenzyme M) 		0.14 g 0.137 0.961
Methanol
(50% v/v) 		20.00 ml 15.469 482.765
Wolin's vitamin solution (10x) 1.00 ml - -
L-Cysteine HCl x H2O* 0.36 g 0.352 2.002
Na2S x 9 H2O* 0.36 g 0.352 1.464
DL-Dithiothreitol* 0.50 g - -
Distilled water 1000.00 ml - -
1 Dissolve ingredients (except bicarbonate, coenzyme M, vitamins, methanol, cysteine and sulfide) and sparge medium with 80% H2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Then 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. Methanol (50% v/v stock solution) and the reducing agents are each autoclaved separately under 100% N2 gas atmosphere as concentrated solutions in tightly closed tubes. Vitamins and coenzyme M are prepared under 100% N2 gas atmosphere and sterilized by filtration. Appropriate volumes of the stock solutions are injected into the sterile medium with hypodermic syringes. Adjust pH of the complete medium to 7.0 - 7.2, if necessary.
2 After inoculation, pressurize culture vessels with sterile 80% H2 and 20% CO2 gas mixture to 1 bar overpressure.

Trace element solution SL-10 #

1000
Solution occurs in 266 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.

Wolin's vitamin solution (10x) #

1000
Solution occurs in 306 other media (first defined in Medium 120).
Compound Amount Unit Conc. [g/L] Conc. [mM]
Biotin 20 mg 0.02 0.082
Folic acid 20 mg 0.02 0.045
Pyridoxine hydrochloride 100 mg 0.1 0.486
Thiamine HCl 50 mg 0.05 0.148
Riboflavin 50 mg 0.05 0.133
Nicotinic acid 50 mg 0.05 0.406
Calcium D-(+)-pantothenate 50 mg 0.05 0.21
Vitamin B12 1 mg 1.0e-3 7.4e-4
p-Aminobenzoic acid 50 mg 0.05 0.365
(DL)-alpha-Lipoic acid 50 mg 0.05 0.242
Distilled water 1000 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: 01.12.22
Source: DSMZ
Taxonomic range:  Archaea
Medium type: Complex medium
Final pH: 7.0 - 7.2
Equipment needed: Autoclave, Filter, 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: 7
Strains with modifications: 4
Methanimicrococcus hongohii DSM 114388

  L-Cysteine HCl x H2O
  Na2S x 9 H2O
  0.5 g DL-Dithiothreitol

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Methanol 15.469 482.765
Na-acetate 2.441 29.761
NaHCO3 2.441 29.062
NaCl 2.197 37.599
Yeast extract 1.953 -
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Cultivation metadata from 7 strains BacDive

phyla 1 5 strains
oxygen anaerobe 2 strains
temperature mesophilic 2 strains