Methanohalobium 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]

Methanohalobium Medium
山东拓普生物工程有限公司 培养基配方 http://www.topbiol.com
NaCl..........................................................................................250.0g
MgSO 4 ·7H 2 O................................................................................4.0g
CaCl 2 ·2H 2 O.................................................................................0.33g
KCl..............................................................................................0.33g
KH 2 PO 4 .......................................................................................0.33g
MgCl 2 ·6H 2 O................................................................................0.33g
NH 4 Cl .........................................................................................0.33g
Resazurin ...................................................................................1.0mg
NaHCO 3  solution ...................................................................100.0mL
Trimethylamine·HCl solution ..................................................20.0mL
Na 2 S·9H 2 O solution.................................................................10.0mL
Yeast extract solution...............................................................10.0mL
Vitamin solution.......................................................................10.0mL
Trace elements solution SL-10 ................................................10.0mL
Na 2 CO 3  solution......................................................................variable
pH 7.4 ± 0.2 at 25°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.
Trimethylamine·HCl Solution:
Composition per 20.0mL:
Trimethylamine·HCl.....................................................................0.2g
Preparation of Trimethylamine·HCl Solution:  Add trimethyl-
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.
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. Mix thoroughly.
Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Yeast Extract Solution:
Composition per 5.0mL:
Yeast extract.............................................................................50.0mg
Preparation of Yeast Extract Solution:  Add yeast extract to dis-
tilled/deionized water and bring volume to 5.0mL. Mix thoroughly.
Sparge with 100% N 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
Lipoic 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 .
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.
Na 2 CO 3 Solution:
Composition per 10.0mL:
Na 2 CO 3 .........................................................................................0.5g
Preparation of Na 2 CO 3 Solution:  Add Na 2 CO 3 to distilled/deion-
ized water and bring volume to 10.0mL. Mix thoroughly. 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 NaHCO 3  solution, trimethyl-
amine·HCl solution, Na 2 S·9H 2 O solution, yeast extract solution, vita-
min solution, trace elements solution SL-10, and Na 2 CO 3  solution, to
distilled/deionized water and bring volume to 840.0mL. Mix thorough-
ly. Autoclave for 15 min at 15 psi pressure–121°C. Anaerobically add
100.0mL of sterile NaHCO 3  solution, 20.0mL of sterile trimethyl-
amine·HCl solution, 10.0mL of sterile Na 2 S·9H 2 O solution, 10.0mL of
sterile yeast extract solution, 10.0mL of sterile vitamin solution, and
10.0mL of sterile trace elements solution SL-10. Mix thoroughly. Add
a sufficient quantity of sterile Na 2 CO 3 solution to bring the pH to 7.4.
Use:  For the cultivation and maintenance of Methanohalobium eves-
tigatus.