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120. METHANOSARCINA MEDIUM

Culture medium recipe

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* Modified for Methanosarcina thermophila DSM 1825
beta

Supplement medium with 5% (v/v) clarified rumen fluid (see medium 1310) and adjust pH to 6.5 - 6.8.

Equipment needed: Hungate tubes Filter Autoclave
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.35 g 0.342 1.962
2 0.23 g 0.225 1.651
3 0.50 g 0.488 9.128
4 0.50 g 0.488 1.981
5 0.25 g 0.244 1.661
6 , might need strain adjustment 2.25 g - -
7
(0.1% w/v in 0.1 N H2SO4)
2.00 ml 0.002 0.007
8 Trace element solution SL-10 1.00 ml - -
9
(OXOID)
2.00 g 1.953 -
10
(BD BBL)
2.00 g 1.953 -
11 2.50 g 2.441 29.761
12
(0.1% w/v)
0.50 ml 4.9e-4 0.002
13 2.00 g 1.953 23.25
14 Vitamin solution 10.00 ml - -
15 10.00 ml 7.715 240.774
16 0.30 g 0.293 1.668
17 0.30 g 0.293 1.22
18 1000.00 ml 974.812 54110.2
19 Clarified rumen fluid , might need strain adjustment
(5% v/v)
1.00 ml - -
20 , might need strain adjustment 5.00 g/l - -
Instructions:
  1. Dissolve ingredients (except bicarbonate, vitamins, methanol, cysteine and sulfide) and sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Then add and dissolve bicarbonate, adjust pH to 6.8 and dispense medium under 80% N2 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 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 6.8 - 7.0, if necessary.
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 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 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
Medium type: Complex medium
Final pH: * 6.5 - 6.8
Equipment needed: Autoclave, Centrifuge, Filter, Hungate tubes

Associated strains

All strains for this medium: 24
Strains with modifications: 24
Methanosarcina thermophila DSM 1825

  6.5 - 6.8

Gas composition

  • 100 % N2

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Distilled water 985.525 54704.9
Methanol 7.715 240.774
Sodium acetate 2.441 29.761
NaHCO3 1.953 23.25
Casitone 1.953 -
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Cultivation metadata from 24 strains BacDive

halophily not 1 strain
pH 8 - 10 1 strain
pH 6 - 8 1 strain
pH 4 - 6 1 strain
phyla 1 24 strains
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