Fumarate Medium (DSMZ Medium 195a)

[所属分类:培养基配方] [发布时间:2021-3-4] [发布人: ] [阅读次数:] [返回]

Fumarate Medium

(DSMZ Medium 195a)

培养基配方

Composition per 991.0mL:
Solution A..............................................................................870.0mL
Solution C..............................................................................100.0mL
Solution D................................................................................10.0mL
Solution E (Vitamin solution)..................................................10.0mL
Solution B (Trace elements solution Sl-10)...............................1.0mL
pH 7.4 ± 0.2 at 25°C
Solution A:
Composition per 870.0mL:
NaCl............................................................................................21.0g
MgCl 2 ·6H 2 O .................................................................................3.1g
Na 2 SO 4 ..........................................................................................3.0g
Na 2 -fumarate.................................................................................2.5g
KCl................................................................................................0.5g
NH 4 Cl ...........................................................................................0.3g
KH 2 PO 4 .........................................................................................0.2g
CaCl 2 ·2H 2 O ................................................................................0.15g
Resazurin ...................................................................................1.0mg
Preparation of Solution A: Add components to distilled/deionized water and bring volume to 870.0mL Mix thoroughly.
Solution B (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 Solution B (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 thoroughly. Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Solution C:
Composition per 100.0mL:
NaHCO 3 ........................................................................................5.0g
Preparation of Solution C: Add NaHCO 3 to distilled/deionized water and bring volume to 100.0mL. Mix thoroughly. Filter sterilize. Flush with 80% N 2  + 20% CO 2 to remove dissolved oxygen.
Solution D:
Composition per 100.0mL:
Resorcinol.....................................................................................1.1g
Preparation of Solution D: Add resorcinol to distilled/deionized water and bring volume to 100.0mL. Mix thoroughly. Sparge with 100% N 2 . Filter sterilize. Use freshly prepared solution.
Solution E (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 ..............................................................................0.10mg
Preparation of Solution E (Vitamin Solution): Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Solution F:
Composition per 10.0mL:
Na 2 S·9H 2 O....................................................................................0.4g
Preparation of Solution F:  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.
Preparation of Medium: Gently heat solution A and bring to boiling. Boil solution A for a few minutes. Cool to room temperature. Gas with 80% N 2  + 20% CO 2 gas mixture to reach a pH below 6. Autoclave for 15 min at 15 psi pressure–121°C. Cool to room temperature. Sequentially add 1.0mL solution B, 100.0mL solution C, 10.0mL solution D, 10.0mL solution E, and 10.0mL solution F. Adjust pH to 7.4 with sodium bicarbonate or sodium carbonate. Distribute anaerobically under 80% N 2  + 20% CO 2 into appropriate vessels. Addition of 10–20mg sodium dithionite per liter from a 5% (w/v) solution, freshly prepared under N 2  and filter-sterilized, may stimulate growth. During growth the culture can be fed with the resorcinol solution.
Use: For the cultivation of Desulfuromusa kysingii, Desulfuromusa bakii, and Desulfuromusa succinoxidans.