Methanosarcina Medium

[所属分类:培养基配方] [发布时间:2021-5-26] [发布人:网站管理员2] [阅读次数:] [返回]

Methanosarcina Medium

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

Composition per liter:

NaCl............................................................................................2.25g
Pancreatic digest of casein............................................................2.0g
Yeast extract..................................................................................2.0g
NaHCO 3 ......................................................................................0.85g
MgSO 4 ·7H 2 O................................................................................0.5g
NH 4 Cl ...........................................................................................0.5g
K 2 HPO 4 ....................................................................................0.348g
CaCl 2 ·2H 2 O................................................................................0.25g
KH 2 PO 4 ....................................................................................0.227g
FeSO 4 ·7H 2 O..............................................................................2.0mg
Resazurin...................................................................................1.0mg
Methanol solution....................................................................10.0mL
Vitamin solution.......................................................................10.0mL
L -Cysteine·HCl·H 2 O solution..................................................10.0mL
Na 2 S·9H 2 O solution.................................................................10.0mL
Trace elements solution SL-10..................................................1.0mL
pH 6.5–6.8 at 25°C
Methanol Solution:
Composition per 10.0mL:
Methanol....................................................................................5.0mL
Preparation of Methanol Solution:  Add methanol to distilled/de-
ionized water and bring volume to 10.0mL. Sparge with 100% N 2 . Au-
toclave for 15 min at 15 psi pressure–121°C.
Vitamin Solution:
Composition per liter:
Pyridoxine·HCl........................................................................10.0mg
Calcium  DL -pantothenate...........................................................5.0mg
Lipoic acid.................................................................................5.0mg
Nicotinic acid.............................................................................5.0mg
p-Aminobenzoic acid.................................................................5.0mg
Riboflavin ..................................................................................5.0mg
Thiamine·HCl ............................................................................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. Filter ster-
ilize. Sparge with 80% N 2 + 20% CO 2 .
L -Cysteine·HCl·H 2 O Solution:
Composition per 10.0mL:
L -Cysteine·HCl·H 2 O.....................................................................0.3g
Preparation of  L -Cysteine·HCl·H 2 O Solution: Add  L -cyste-
ine·HCl·H 2 O to distilled/deionized water and bring volume to 10.0mL.
Mix thoroughly. Autoclave under 100% N 2 for 15 min at 15 psi pres-
sure–121°C.
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 distilled/
deionized water and bring volume to 10.0mL. Mix thoroughly. Sparge
with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Trace Elements Solution SL-10:
Composition per liter:
FeCl 2 ·4H 2 O...................................................................................1.5g
CoCl 2 ·6H 2 O ...........................................................................190.0mg
MnCl 2 ·4H 2 O...........................................................................100.0mg
ZnCl 2 ........................................................................................70.0mg
Na 2 MoO 4 ·2H 2 O .......................................................................36.0mg
NiCl 2 ·6H 2 O..............................................................................24.0mg
H 3 BO 3 ........................................................................................6.0mg
CuCl 2 ·2H 2 O ...............................................................................2.0mg
HCl (25% solution)..................................................................10.0mL
Preparation of Trace Elements Solution SL-10: Add FeCl 2 ·4H 2 O
to 10.0mL of HCl solution. Mix thoroughly. Add distilled/deionized
water and bring volume to 1.0L. Add remaining components. Mix thor-
oughly. Sparge with 100% N 2 .
Preparation of Medium: Add components, except methanol solution,
vitamin solution,  L -cysteine·HCl·H 2 O solution, and Na 2 S·9H 2 O solution,
to distilled/deionized water and bring volume to 960.0mL. Mix thorough-
ly. Sparge under 80% N 2 + 20% CO 2 for 3–4 min. Autoclave for 15 min at
15 psi pressure–121°C. Aseptically and anaerobically add 10.0mL of ster-
ile methanol, 10.0mL of sterile vitamin solution, 10.0mL of sterile  L -
cysteine·HCl·H 2 O solution, and 10.0mL of sterile Na 2 S·9H 2 O solution.
Mix thoroughly. Aseptically and anaerobically distribute into sterile
screw-cappped bottles under 80% N 2 + 20% CO 2 .
Use: For the cultivation and maintenance of Methanosarcina species.