Basal Medium (DSMZ Medium 1001)

[所属分类:培养基配方] [发布时间:2019-12-6] [发布人:] [阅读次数:] [返回]

山东拓普生物工程有限公司

Shandong Tuopu Biol-Engineering Co.,Ltd

Basal Medium
(DSMZ Medium 1001)

培养基配方
Composition per liter:
NaHCO3 ........................................................2.5g
NH4Cl ........................................................0.25g
NaH2PO4 .......................................................0.6g
KCl............................................................1.0g
Iron nitrilotriacetic acid solution .........................20.0mL
Vitamin mix..................................................10.0mL
Mineral mix .................................................10.0mL
Sodium acetate solution......................................10.0mL
pH 6.9 ± 0.2 at 25°C
Mineral Mix:
Composition per liter:
MgSO4 ·7H2O................................................3.0g
Nitrilotriacetic acid.......................................1.5g
NaCl........................................................1.0g
MnSO4 ·2H2O................................................0.5g
ZnCl2 .....................................................0.13g
CoCl2 ·6H2O................................................0.1g
CaCl2 ·2H2O ...............................................0.1g
FeSO4 ·7H2O................................................0.1g
Na2MoO4 ·4H2O............................................0.025g
NaWO4 ·2H2O .............................................0.025g
NiCl2 ·6H2O..............................................0.024g
CuSO4 ·5H2O..............................................0.01g
KAl(SO4)2 ·12H2O.........................................0.01g
H3BO3 ....................................................0.01g
Preparation of Mineral Mix:  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 Mix:
Composition per liter:
Pyridoxine-HCl...........................................10.0mg
Thiamine-HCl·2H2O........................................5.0mg
Riboflavin................................................5.0mg
Nicotinic acid............................................5.0mg
D -Ca-pantothenate .......................................5.0mg
p-Aminobenzoic acid.......................................5.0mg
Thioctic acid ............................................5.0mg
Biotin....................................................2.0mg
Folic acid ...............................................2.0mg
Vitamin B 12 .............................................0.1mg
Preparation of Vitamin Mix:  Add components to distilled/deion-
ized water and bring volume to 1.0L. Mix thoroughly. Sparge with 80%
H 2 + 20% CO 2 . Filter sterilize.
Sodium Acetate Solution:
Composition per 100.0mL:
Sodium acetate.................................................13.6g
Preparation of Sodium Acetate Solution:  Add sodium acetate to
distilled/deionized water and bring volume to 80.0mL with distilled/
deionized water. Mix thoroughly. Bring volume to 100.0mL with dis-
tilled/deionized water. Sparge with 100% N 2 for 45 min. Seal in bottle.
Autoclave for 15 min at 15 psi pressure–121°C.
Iron Nitriloacetic Acid Solution:
Composition per 100.0mL:
FeCl3 ·6H2O..................................................13.5g
Sodium nitrilotriacetic acid (NTA)............................12.8g
NaHCO3 ........................................................8.2g
Preparation of Iron Nitriloacetic Acid:  Add NaHCO3 to dis-
tilled/deionized water and bring volume to 70.0mL with distilled/de-
ionized water. Mix thoroughly. Add NTA. Mix thoroughly. Add
FeCl3 ·6H2O. Adjust pH to 6.5 using 10N NaOH. Bring volume to
100.0mL with distilled/deionized water. Stir for about 15 minutes to al-
low components to go into solution. Sparge with 100% N2 for 45 min.
Filter sterilize. Aseptically and anoxically dispense into sterile serum
bottles.
Preparation of Medium: Add components, except iron nitrilo-
acetic acid solution and sodium acetate solution, to distilled/deionized
water and bring volume to 1.0L. Mix thoroughly. Bubble the medium
with 80:20 N2 :CO2 (final pH should be 6.8 to 7.0). Approximately
10.0mL of media (anaerobic culture tube) should be gassed for 5 min
in the aqueous phase (bubbled) and the headspace gassed for 1 min e
prior to sealing the container. Autoclave for 15 min at 15 psi pressure–
121°C. Add electron donor (acetate-final conc. of 10mM-) and electron
acceptor (Fe(III)NTA (final conc. of 10mM), from sterile, anaerobic
stock solutions using a sterile syringe and needle flushed with anaero-
bic gas. This medium should not be exposed to direct sunlight!
Use:  For the cultivation of a Rhodoferax ferrireducens.