KCTC Media No. 1136 : Methanobacterium medium

조성	용량	단위
KH2PO4	0.5	g
MgSO47H2O	0.4	g
NaCl	0.4	g
NH4Cl	0.4	g
CaCl22H2O	0.05	g
FeSO47H2O solution(0.1% w/v in 0.1N H2SO4)	2.0	ml
조성 용량 단위 H2SO4 (0.1 N) 1000 ml FeSO4· 7H2O 1.0 g The ferrous sulfate solution is not stable and should be freshly prepared.
Yeast extract(OXOID)	1.0	g
Na-acetate	1.0	g
Na-formate	2.0	g
NaHCO3	4	g
Resazurin	1.0	mg
L-Cystein-HCl·H2O	0.5	g
Na2S·9H2O	0.5	g
D·W	930	ml
Trace element solution SL-10	1.0	ml
조성 용량 단위 HCl (25%) 10.0 ml FeCl2·4H2O 1.5 g ZnCl2 70 mg MnCl2·4H2O 100 mg H3BO3 6 mg CoCl2·6H2O 190 mg CuCl2·2H2O 2.0 mg NiCl·6H2O 24 mg Na2MoO4·2H2O 36 mg D·w 990 ml First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml.
Sludge fluid	50	ml
조성 용량 단위 Yeast extract 4 g Sludge 1000 ml
Fatty acid mixture	20.0	ml
조성 용량 단위 Isobutyric acid 23 ml Valeric acid 27 ml Isovaleric acid 27 ml 2-Methylbutyric acid 27 ml D·W 896 ml Adjust pH to 7.5 with concentrated NaOH.

** ph : 6.8-7.0

** memo : -

** 추가조치 사항 : 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, then dispense medium under 80% H2 and 20% CO2 gas atmosphere into
anoxic Hungate-type tubes 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.
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:
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 room temperature in the
dark.