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DSMZ

DSMZ 404 . METHANOSPHAERA CUNICULI MEDIUM

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

Culture medium recipe

Vol.
×

Main sol. 404 #

ml
Equipment needed: Hungate tubes Filter Autoclave Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
Mineral solution 1 40.0 ml - -
Minerals solution 40.0 ml - -
Fe(NH4)2(SO4)2 x 7 H2O
(0.1% w/v) 		2.0 ml 0.002 0.005
NiCl2 x 6 H2O
(0.1% w/v) 		0.2 ml 2.0e-4 0.002
Modified Wolin's mineral solution 10.0 ml - -
Na-acetate x 3 H2O 3.0 g 2.988 -
Yeast extract
(OXOID) 		1.0 g 0.996 -
Trypticase peptone
(BD BBL) 		1.0 g 0.996 -
Sodium resazurin
(0.1% w/v) 		0.5 ml 5.0e-4 0.002
NaHCO3 3.0 g 2.988 35.569
Methanol
(50% v/v) 		10.0 ml 7.888 246.191
Wolin's vitamin solution (10x) 1.0 ml - -
L-Cysteine HCl x H2O 0.3 g 0.299 1.701
Na2S x 9 H2O 0.3 g 0.299 1.244
Distilled water 900.0 ml - -
1 Dissolve ingredients (except bicarbonate, methanol, vitamins, cysteine and sulfide), then sparge medium with 80% H2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Add and dissolve bicarbonate and adjust pH to 7.0, then distribute under 80% H2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials to 30% of their volume and autoclave. After sterilization add methanol (50% v/v), vitamins, cysteine and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas atmosphere. Vitamins are sterilized by filtration. Adjust pH of complete medium to 7.0 - 7.2.
2 After inoculation, pressurize culture bottles with sterile 80% H2 and 20% CO2 gas mixture to 2 bar overpressure.

Mineral solution 1 #

1000
Solution occurs in 4 other media (first defined in Medium 203).
Compound Amount Unit Conc. [g/L] Conc. [mM]
K2HPO4 6 g 6 34.448
Distilled water 1000 ml - -

Minerals solution #

1000
Solution occurs in 2 other media (first defined in Medium 317).
Compound Amount Unit Conc. [g/L] Conc. [mM]
KH2PO4 6.00 g 6 44.09
(NH4)2SO4 6.00 g 6 45.406
NaCl 12.00 g 12 205.339
MgSO4 x 7 H2O 2.60 g 2.6 10.549
CaCl2 x 2 H2O 0.16 g 0.16 1.088
Distilled water 1000.00 ml - -

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.

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: 02.12.22
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: 1

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Methanol 7.888 246.191
NaHCO3 2.988 35.569
Na-acetate x 3 H2O 2.988 -
Yeast extract 0.996 -
Trypticase peptone 0.996 -
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Cultivation metadata from 1 strains BacDive

phyla 1 1 strain
oxygen anaerobe 1 strain
temperature mesophilic 1 strain