Methanohalophilus halophilus Medium

[所属分类:培养基配方] [发布时间:2021-5-25] [发布人:网站管理员2] [阅读次数:] [返回]

Methanohalophilus halophilus Medium

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

Composition per 1011.0mL:

NaCl............................................................................................70.0g
KCl..............................................................................................0.33g
KH 2 PO 4 .......................................................................................0.33g
NH 4 Cl .........................................................................................0.33g
Yeast extract................................................................................0.05g
K 2 SO 4 ..........................................................................................0.01g
Resazurin ...................................................................................1.0mg
Methylamine·HCl solution.......................................................20.0mL
MgCl 2 /CaCl 2 solution..............................................................10.0mL
NaHCO 3  solution .....................................................................10.0mL
Vitamin solution.......................................................................10.0mL
L -Cysteine·HCl solution...........................................................10.0mL
Na 2 S·9H 2 O solution.................................................................10.0mL
Trace elements solution SL-10 ..................................................1.0mL
pH 7.2 ± 0.2 at 25°C
MgCl 2 /CaCl 2 Solution:
Composition per 10.0mL:
MgCl 2 ·6H 2 O................................................................................0.33g
CaCl 2 ·2H 2 O.................................................................................0.33g
Preparation of MgCl 2 /CaCl 2 Solution:  Add components to dis-
tilled/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.
NaHCO 3 Solution:
Composition per 10.0mL:
NaHCO 3 ........................................................................................5.0g
Preparation of NaHCO 3 Solution:  Add NaHCO 3 to distilled/de-
ionized water and bring volume to 10.0mL. Mix thoroughly. Sparge
with 80% N 2 + 20% CO 2 . Autoclave for 15 min at 15 psi pressure–
121°C.
Vitamin Solution:
Composition per liter:
Pyridoxine·HCl........................................................................10.0mg
Calcium  DL -pantothenate...........................................................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% H 2 + 20% CO 2 .
Methylamine·HCl Solution:
Composition per 20.0mL:
Methylamine·HCl .........................................................................5.0g
Preparation of Methylamine·HCl Solution:  Add methyl-
amine·HCl to distilled/deionized water and bring volume to 20.0mL.
Mix thoroughly. Sparge with 100% N 2 . Autoclave for 15 min at 15 psi
pressure–121°C.
L -Cysteine·HCl Solution:
Composition per 10.0mL:
L -Cysteine·HCl..............................................................................0.2g
Preparation of  L -Cysteine·HCl Solution: Add  L -cysteine·HCl to
distilled/deionized water and bring volume to 10.0mL. Mix thorough-
ly. Autoclave under 100% N 2 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....................................................................................0.2g
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. 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 . Autoclave for 15 min at 15 psi pressure–
121°C.
Preparation of Medium: Prepare and dispense medium under 80%
H 2 + 20% CO 2 . Add components, except MgCl 2 /CaCl 2 solution,
NaHCO 3  solution, vitamin solution, methylamine·HCl solution,  L -
cysteine·HCl solution, and Na 2 S·9H 2 O solution, to distilled/deionized
water and bring volume to 840.0mL. Mix thoroughly. Adjust pH to
6.9–7.0. Sparge with 80% H 2 + 20% CO 2 . Autoclave for 15 min at 15
psi pressure–121°C. Anaerobically add 10.0mL of sterile MgCl 2 /CaCl 2
solution, 10.0mL of sterile NaHCO 3  solution, 10.0mL of sterile vita-
min solution, 20.0mL of sterile methylamine·HCl solution, 10.0mL of
sterile  L -cysteine·HCl solution, and 10.0mL of sterile Na 2 S·9H 2 O solu-
tion. Mix thoroughly.
Use: For the cultivation and maintenance of Methanohalophilus halo-
philus.