Methanobacterium subterraneum Medium (DSMZ Medium 814)

[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 subterraneum Medium
(DSMZ Medium 814)

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

Composition per liter:

Na-formate....................................................................................3.2g
Na-acetate .....................................................................................1.0g
Tryptone........................................................................................1.0g
K 2 HPO 4 .........................................................................................0.5g
NaCl............................................................................................0.45g
NH 4 Cl ...........................................................................................0.4g
MgCl 2 ·6H 2 O................................................................................0.03g
FeSO 4 ·7H 2 O..............................................................................0.003g
Resazurin ...................................................................................0.5mg
Trace elements solution ...........................................................10.0mL
Cysteine solution......................................................................10.0mL
Na 2 S·9H 2 O solution.................................................................10.0mL
NaHCO 3  solution .....................................................................10.0mL
Seven vitamin solution...............................................................1.0mL
pH 8.3 ± 0.2 at 25°C
NaHCO 3 Solution:
Composition per 10.0mL:
NaHCO 3 ........................................................................................2.0g
Preparation of NaHCO 3 Solution:  Add NaHCO 3 to distilled/de-
ionized water and bring volume to 10.0mL. Mix thoroughly. Sparge
with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C. Cool to
25°C. Must be prepared freshly.
Cysteine Solution:
Composition per 10.0mL:
L -Cysteine-HCl·H 2 O...................................................................0.25g
Preparation of Cysteine Solution:  Add  L -cysteine·HCl·H 2 O to
distilled/deionized water and bring volume to 10.0mL. Mix thorough-
ly. Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–
121°C.
Na 2 S·9H 2 O Solution:
Composition per 10.0mL:
Na 2 S·9H 2 O..................................................................................0.25g
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.
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.
Seven Vitamin Solution:
Composition per liter:
Pyridoxine hydrochloride......................................................300.0mg
Thiamine-HCl·2H 2 O..............................................................200.0mg
Nicotinic acid.........................................................................200.0mg
Vitamin B 12 ............................................................................100.0mg
Calcium pantothenate ............................................................100.0mg
p-Aminobenzoic acid...............................................................80.0mg
D (+)-Biotin...............................................................................20.0mg
Preparation of Seven Vitamin Solution: Add components to dis-
tilled/deionized water and bring volume to 1.0L. Sparge with 100% N 2 .
Mix thoroughly. Filter sterilize.
Preparation of Medium: Prepare and dispense medium under 100%
N 2 . Add components, except Na 2 S·9H 2 O solution, seven vitamin solu-
tion, cysteine solution, and NaHCO 3  solution, to distilled/deionized wa-
ter and bring volume to 969.0mL. Mix thoroughly. Flush medium with
N 2 for 5 min. Adjust medium pH to 8.3. Autoclave for 15 min at 15 psi
pressure–121°C. Aseptically and anaerobically add 1.0mL seven vita-
min mix, 10.0mL cysteine solution, 10.0mL Na 2 S·9H 2 O, and 10.0mL
NaHCO 3  solution. Mix thoroughly. Adjust pH to 8.3. Aseptically and
anaerobically distribute into sterile tubes or bottles.
Use:  For the cultivation of Methanobacterium subterraneum.