R8 Medium (DSMZ Medium 912)

[所属分类:培养基配方] [发布时间:2021-7-30] [发布人:网站管理员2] [阅读次数:] [返回]

R8 Medium
(DSMZ Medium 912)

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

Composition per liter:

NaHCO 3 ......................................................................................2.52g
MOPS ...........................................................................................2.1g
NaCl..............................................................................................1.0g
Glucose.........................................................................................0.9g
MgCl 2 ·6H 2 O .................................................................................0.5g
Na 2 S·9H 2 O....................................................................................0.6g
Cysteine-HCl ................................................................................0.4g
NH 4 Cl ...........................................................................................0.3g
KCl................................................................................................0.3g
K 2 HPO 4 .......................................................................................0.25g
KH 2 PO 4 .........................................................................................0.2g
Yeast extract................................................................................0.19g
Peptone........................................................................................0.19g
CaCl 2 ·2H 2 O...............................................................................0.015g
Resazurin ...................................................................................0.5mg
Rumen fluid .............................................................................50.0mL
Na-pantothenate solution.........................................................10.0mL
Trace elements solution SL-10 ..................................................1.0mL
pH 7.2 ± 0.2 at 25°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 .
Rumen Fluid:
Composition per 50.0mL:
Rumen fluid, clarified..............................................................50.0mL
Preparation of Rumen Fluid: Sparge clarified rumen fluid with
100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Na-Pantothenate Solution:
Composition per 10.0mL:
Na-pantothenate............................................................................0.1g
Preparation of Na-Pantothenate Solution: Add Na-pantothen-
ate to distilled/deionized water and bring volume to 10.0mL. Mix thor-
oughly. Sparge with 100% N 2 . Filter sterilize.
Preparation of Medium: Prepare and dispense medium under 80%
N 2 + 20% CO 2 gas atmosphere. Add components, except NaHCO 3 , pan-
tothenate solution, Na 2 S·9H 2 O, cysteine-HCl, and rumen fluid, to dis-
tilled/deionized water and bring volume to 940.0mL. Mix thoroughly.
Gently heat and bring to boiling. Boil for 3 min. Cool to room tempera-
ture while sparging with 80% N 2 + 20% CO 2 . Add solid bicarbonate, so-
dium sulfide, and cysteine-HCl. Adjust pH to 7.2. Distribute under 80%
N 2 + 20% CO 2  atmosphere into anaerobe tubes or bottles. Autoclave for
15 min at 15 psi pressure–121°C. Aseptically and anaerobically add, per
liter of medium, 50.0mL rumen fluid and 10.0mL Na-pantothenate so-
lution. Mix thoroughly. The final pH of the medium should be 7.2.
Use:  For the cultivation of Spirochaeta spp.