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DSMZ

DSMZ 161 . METHANOFOLLIS MEDIUM

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

Culture medium recipe

Vol.
×

Main sol. 161 #

ml
Equipment needed: Hungate tubes Autoclave Filter Gassing station Centrifuge
Compound Amount Unit Conc. [g/L] Conc. [mM]
Clarified rumen fluid 300.00 ml - -
K2HPO4 0.30 g 0.3 1.721
KH2PO4 0.30 g 0.3 2.202
(NH4)2SO4 0.30 g 0.3 2.268
NaCl 0.60 g 0.599 10.257
MgSO4 x 7 H2O 0.13 g 0.13 0.527
CaCl2 x 2 H2O
(0.1% w/v) 		8.00 ml 0.008 0.054
Modified Wolin's mineral solution 10.00 ml - -
FeSO4 x 7 H2O
(0.1% w/v in 0.1 N H2SO4) 		2.00 ml 0.002 0.007
Yeast extract
(DIFCO) 		1.00 g 0.999 -
Trypticase
(BBL) 		1.00 g 0.999 -
Fatty acid mixture 20.00 ml - -
Sodium resazurin
(0.1% w/v) 		0.50 ml 5.0e-4 0.002
NaHCO3 2.00 g 1.998 23.784
Wolin's vitamin solution (10x) 1.00 ml - -
L-Cysteine HCl x H2O 0.50 g 0.5 2.844
Na2S x 9 H2O 0.50 g 0.5 2.08
Distilled water 660.00 ml - -
1 Dissolve ingredients (except bicarbonate, vitamins, cysteine and sulfide) and sparge medium for 30 - 45 min with 80% H2 and 20% CO2 gas mixture to make it anoxic. Add and dissolve bicarbonate and adjust pH to 6.8, then distribute under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After sterilization add cysteine and sulfide from sterile anoxic stock solutions autoclaved under 100% N2 gas atmosphere. Vitamins are prepared under 100% N2 gas atmosphere and sterilized by filtration. Adjust pH of complete medium to 6.5. For incubation use sterile 80% H2 and 20% CO2 gas mixture at two atmospheres of pressure.

Clarified rumen fluid #

1000
Solution occurs in 28 other media (first defined in Medium 1310).
1 Rumen fluid from cow or sheep (obtained from fistulated animals or abattoir refuse) is filtered through muslin, autoclaved at 121°C for 15 min and 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.

Modified Wolin's mineral solution #

1000
Solution occurs in 166 other media (first defined in Medium 141).
Compound Amount Unit Conc. [g/L] Conc. [mM]
Nitrilotriacetic acid 1.50 g 1.5 7.848
MgSO4 x 7 H2O 3.00 g 3 12.172
MnSO4 x H2O 0.50 g 0.5 2.958
NaCl 1.00 g 1 17.112
FeSO4 x 7 H2O 0.10 g 0.1 0.36
CoSO4 x 7 H2O 0.18 g 0.18 1.161
CaCl2 x 2 H2O 0.10 g 0.1 0.68
ZnSO4 x 7 H2O 0.18 g 0.18 0.626
CuSO4 x 5 H2O 0.01 g 0.01 0.04
AlK(SO4)2 x 12 H2O 0.02 g 0.02 0.042
H3BO3 0.01 g 0.01 0.162
Na2MoO4 x 2 H2O 0.01 g 0.01 0.041
NiCl2 x 6 H2O 0.03 g 0.03 0.231
Na2SeO3 x 5 H2O 0.30 mg 3.0e-4 0.001
Na2WO4 x 2 H2O 0.40 mg 4.0e-4 0.001
Distilled water 1000.00 ml - -
1 First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH.

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.

Wolin's vitamin solution (10x) #

1000
Solution occurs in 305 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 - -

Metadata

Last modified: 26.02.24
Source: DSMZ
Synonyms: METHANOMICROBIUM MOBILE MEDIUM
Taxonomic range:  Archaea
Medium type: Complex medium
Final pH: 6.5
Equipment needed: Autoclave, Centrifuge, 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: 1

Molecular composition

Compound Conc. [g/L] Conc. [mM]
NaHCO3 1.998 23.784
Yeast extract 0.999 -
Trypticase 0.999 -
L-Cysteine HCl x H2O 0.5 2.844
Na2S x 9 H2O 0.5 2.08
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Cultivation metadata from 1 strains BacDive

phyla 1 1 strain
temperature mesophilic 1 strain