Methanobacterium II Medium (DSMZ Medium 825)

[所属分类:培养基配方] [发布时间:2021-5-24] [发布人:网站管理员2] [阅读次数:] [返回]

Methanobacterium II Medium
(DSMZ Medium 825)

山东拓普生物工程有限公司 培养基配方 http://www.topbiol.com

Composition per 1060.0mL:

Yeast extract..................................................................................1.0g
NH 4 Cl ...........................................................................................1.0g
NaCl..............................................................................................0.6g
Cysteine-HCl·H 2 O........................................................................0.5g
Sodium acetate..............................................................................0.5g
K 2 HPO 4 .........................................................................................0.3g
KH 2 PO 4 .........................................................................................0.3g
MgCl 2 ·6H 2 O..................................................................................0.2g
CaCl 2 ·2H 2 O...................................................................................0.1g
KCl................................................................................................0.1g
Resazurin ...................................................................................0.5mg
NaHCO 3 solution.....................................................................40.0mL
Trace elements solution ...........................................................10.0mL
Vitamin solution.......................................................................10.0mL
Na 2 S·9H 2 O solution.................................................................10.0mL
Methanol solution....................................................................10.0mL
pH 7.0 ± 0.2 at 25°C
Methanol Solution:
Composition per 100.0mL:
Methanol..................................................................................15.0mL
Preparation of Methanol Solution:  Add methanol to distilled/de-
ionized water and bring volume to 100.0mL. Mix thoroughly. Sparge
with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Trace Elements Solution:
Composition per liter:
MgSO 4 ·7H 2 O................................................................................3.0g
Nitrilotriacetic acid.......................................................................1.5g
NaCl..............................................................................................1.0g
MnSO 4 ·2H 2 O................................................................................0.5g
CoSO 4 ·7H 2 O...............................................................................0.18g
ZnSO 4 ·7H 2 O...............................................................................0.18g
CaCl 2 ·2H 2 O ..................................................................................0.1g
FeSO 4 ·7H 2 O..................................................................................0.1g
NiCl 2 ·6H 2 O...............................................................................0.025g
KAl(SO 4 ) 2 ·12H 2 O.......................................................................0.02g
H 3 BO 3 .........................................................................................0.01g
Na 2 MoO 4 ·4H 2 O..........................................................................0.01g
CuSO 4 ·5H 2 O...............................................................................0.01g
Na 2 SeO 3 ·5H 2 O...........................................................................0.3mg
Preparation of Trace Elements Solution:  Add nitrilotriacetic
acid to 500.0mL of distilled/deionized water. Dissolve by adjusting pH
to 6.5 with KOH. Add remaining components. Add distilled/deionized
water to 1.0L. Mix thoroughly.
Vitamin Solution:
Composition per liter:
Pyridoxine-HCl........................................................................10.0mg
Thiamine-HCl·2H 2 O..................................................................5.0mg
Riboflavin..................................................................................5.0mg
Nicotinic acid.............................................................................5.0mg
D -Ca-pantothenate .....................................................................5.0mg
p-Aminobenzoic acid.................................................................5.0mg
Lipoic acid.................................................................................5.0mg
Biotin.........................................................................................2.0mg
Folic acid ...................................................................................2.0mg
Vitamin B 12 ................................................................................0.1mg
Preparation of Vitamin Solution:  Add components to distilled/
deionized water and bring volume to 1.0L. Mix thoroughly. Sparge
with 80% H 2 + 20% CO 2 . Filter sterilize.
Na 2 S·9H 2 O Solution:
Composition per 10.0mL:
Na 2 S·9H 2 O....................................................................................0.3g
Preparation of Na 2 S·9H 2 O Solution:  Add Na 2 S·9H 2 O to dis-
tilled/deionized water and bring volume to 10.0mL. Sparge with N 2 .
Autoclave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Store an-
aerobically.
NaHCO 3 Solution:
Composition per 100.0mL:
NaHCO 3 ......................................................................................10.0g
Preparation of NaHCO 3 Solution:  Add NaHCO 3 to distilled/de-
ionized water and bring volume to 100.0mL. Mix thoroughly. Auto-
clave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Must be
prepared freshly.
Preparation of Medium: Prepare and dispense medium under 80%
N 2 + 20% CO 2  gas mixture. Add components, except methanol solu-
tion, NaHCO 3 solution, and Na 2 S·9H 2 O solution, to 1.0L distilled/de-
ionized water. Mix thoroughly. Sparge with 80% N 2 + 20% CO 2 .
Dispense 5.0mL aliquots into Hungate tubes under 80% H 2 + 20% CO 2
gas mixture. Autoclave for 15 min at 15 psi pressure–121°C. Prior to
use inject, for each 5.0mL medium, 0.2mL sterile NaHCO 3 solution,
0.05mL sterile Na 2 S·9H 2 O solution, and 0.05mL sterile methanol so-
lution. The pH should be 7.0.
Use:  For the cultivation of Methanosarcina barkeri DSM 10131 and
Methanosarcina mazei=Methanococcus mazei (Methanosarcina fri-
sia) DSM 10132.