Anaerobic Cholesterol Medium (DSMZ Medium 858)

[所属分类:培养基配方] [发布时间:2019-10-14] [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

Anaerobic Cholesterol Medium
(DSMZ Medium 858)
Composition per 1010mL:
Cholesterol....................................................................................0.5g
Solution A..............................................................................900.0mL
Solution B................................................................................50.0mL
Solution F (NaHCO 3 solution).................................................50.0mL
Solution D (Vitamin solution)....................................................5.0mL
Solution C (Trace elements solution SL-10)..............................2.5mL
Solution E (Selenite tungstate solution).....................................2.5mL
pH 7.0 ± 0.2 at 25°C
Solution A:
Composition per 200mL:
NaNO 3 .........................................................................................0.16g
MgSO 4 ·7H 2 O..............................................................................0.10g
NH 4 Cl .........................................................................................0.10g
CaCl 2 ...........................................................................................0.02g
Preparation of Solution A: Add components to distilled/deionized
water and bring volume to 200.0mL. Sparge with 100% N 2 . Mix thor-
oughly.
Solution B:
Composition per 100.0mL:
KH 2 PO 4 .........................................................................................1.0g
Preparation of Solution B: Add KH 2 PO 4 to distilled/deionized
water and bring volume to 100.0mL. Sparge with 80% N 2 + 20% CO 2 .
Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C.
Solution 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 Solution C (Trace Elements Solution SL–10):
Add FeCl 2 ·4H 2 O to 10.0mL of HCl solution. Mix thoroughly. Add dis-
tilled/deionized water and bring volume to 1.0L. Add remaining com-
ponents. Mix thoroughly. Sparge with 80% N 2 + 20% CO 2 . Autoclave
for 15 min at 15 psi pressure–121°C.
Solution D (Vitamin Solution):
Composition per liter:
Vitamin B 12 ..............................................................................50.0mg
Pantothenic acid.......................................................................50.0mg
Riboflavin................................................................................50.0mg
Alpha-lipoic acid .....................................................................50.0mg
p-Aminobenzoic acid...............................................................50.0mg
Thiamine-HCl·2H 2 O................................................................50.0mg
Nicotinic acid...........................................................................25.0mg
Nicotine amide.........................................................................25.0mg
Biotin.......................................................................................20.0mg
Folic acid .................................................................................20.0mg
Pyridoxamine-HCl...................................................................10.0mg
Preparation of Solution D (Vitamin Solution): Add components
to distilled/deionized water and bring volume to 1.0L. Sparge with
100% N 2 . Mix thoroughly. Filter sterilize.
Solution E (Selenite Tungstate Solution):
Composition per liter:
NaOH............................................................................................0.5g
Na 2 WO 4 ·2H 2 O...........................................................................4.0mg
Na 2 SeO 3 ·5H 2 O...........................................................................3.0mg
Preparation of Solution E (Selenite Tungstate Solution): Add
components to distilled/deionized water and bring volume to 1.0L. Mix
thoroughly. Sparge with 100% N 2 . Filter sterilize.
Solution F (NaHCO 3 Solution):
Composition per 100.0mL:
NaHCO 3 ........................................................................................8.4g
Preparation of Solution F (NaHCO 3 Solution):  Add NaHCO 3
to distilled/deionized water and bring volume to 100.0mL. Mix thor-
oughly. Sparge with 80% N 2 + 20% CO 2 . Autoclave for 15 min at 15
psi pressure–121°C.
Preparation of Medium: Distribute solution A into anaerobic
tubes or bottles. Distribute appropriate amounts of cholesterol, 5.0mg
cholesterol per 10.0mL solution A. Sparge with 80% N 2 + 20% CO 2 .
Autoclave for 15 min at 15 psi pressure–121°C. Cool to room temper-
ature. Add to 9.0mL solution A: 500μl sterile solution B, 500μl sterile
solution F, 25μl sterile solution C, 25μl sterile solution E, and 50μl
sterile solution D. Adjust pH to 7.0.
Use: For the cultivation of unclassified bacterium DSM 12783 and
Sterolibacterium denitrificans DSM 13999 = ATCC BAA-354.