Methanococcus sp. Medium (DSMZ Medium 141a)

[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]

Methanococcus sp. Medium
(DSMZ Medium 141a)

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

Composition per 1040.0mL:

NaCl............................................................................................18.0g
MgCl 2 ·6H 2 O .................................................................................4.0g
MgSO 4 ·7H 2 O..............................................................................3.45g
Na-acetate.....................................................................................1.0g
KCl............................................................................................0.335g
NH 4 Cl .........................................................................................0.25g
CaCl 2 ·2H 2 O ................................................................................0.14g
K 2 HPO 4 .......................................................................................0.14g
Fe(NH 4 ) 2 (SO 4 ) 2 ·7H 2 O...............................................................2.0mg
Resazurin...................................................................................1.0mg
Trace elements solution...........................................................10.0mL
Vitamin solution.......................................................................10.0mL
L -Cysteine solution..................................................................10.0mL
Na 2 S·9H 2 O solution.................................................................10.0mL
pH 7.2 ± 0.2 at 25°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....................................................................................0.5g
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.
L -Cysteine Solution:
Composition per 10.0mL:
L -Cysteine·HCl·H 2 O .....................................................................0.5g
Preparation of  L -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.
Preparation of Medium: Prepare and dispense medium under an
oxygen-free 80% H 2 + 20% CO 2  gas mixture. Add components, except
vitamin solution,  L -cysteine solution, and Na 2 S·9H 2 O solution, to dis-
tilled/deionized water and bring volume to 1.0L. Mix thoroughly.
Sparge with 80% H 2 + 20% CO 2 . Autoclave for 15 min at 15 psi pres-
sure–121°C. Cool to 25°C while sparging with 80% H 2 + 20% CO 2 .
Aseptically and anaerobically add 10.0mL vitamin solution, 10.0mL of
sterile  L -cysteine solution, and 10.0mL of sterile Na 2 S·9H 2 O solution.
Mix thoroughly. Aseptically and anaerobically distribute into sterile
tubes or flasks. Alternately the medium can be distributed to tubes un-
der anaerobic conditions and autoclaved in tubes prior to addition of
substrate solution, vitamin solution, and Na 2 S·9H 2 O solution. Appro-
priate amounts of these solutions can then be added to each tube by sy-
ringes to yield the desired concentrations. After inoculation, pressurize
culture vessels to 2 bar 80% H 2 + 20% CO 2 overpressure.
Use:  For the cultivation of Methanococcus sp.