E Medium for Anaerobes with Filtered Rumen Fluid and 0.1% Cellobiose

[所属分类:培养基配方] [发布时间:2020-7-1] [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

E Medium for Anaerobes with Filtered Rumen Fluid
and 0.1% Cellobiose

培养基配方
Composition per 100.0mL:
Cellobiose....................................................................0.1g
Glucose......................................................................0.05g
L- Cysteine·HCl·H2O........................................................0.05g
Maltose .....................................................................0.05g
(NH4)2SO4 ...................................................................0.05g
Peptone .....................................................................0.05g
Soluble starch...............................................................0.05g
Yeast extract................................................................0.05g
Salts solution..............................................................50.0mL
Rumen fluid, filtered.......................................................30.0mL
Resazurin solution .........................................................0.4mL
pH 7.0 ± 0.2 at 25°C
Salts Solution:
Composition per liter:
NaHCO3 .....................................................................10.0g
NaCl.........................................................................2.0g
K2HPO4 ......................................................................1.0g
KH2PO4 ......................................................................1.0g
CaCl2 (anhydrous) ...........................................................0.2g
MgSO4 .......................................................................0.2g
Preparation of Salts Solution:  Add CaCl2 and MgSO 4 to approx-
imately 300.0mL of distilled/deionized water. Mix thoroughly. Bring
volume to 800.0mL with distilled/deionized water. Add remaining
components. Mix thoroughly. Bring volume to 1.0L with distilled/de-
ionized water. Mix thoroughly. Store at 4°C.
Rumen Fluid:
Composition per 100.0mL:
Rumen fluid.............................................................100.0mL
Preparation of Rumen Fluid:  Obtain the rumen contents from a
cow that has been fed an alfalfa-hay ration. Filter rumen contents
through two layers of cheesecloth. Store under 100% CO2 in the refrig-
erator. The particulate material will settle out. Use only the supernatant
liquid. Filter through a 0.20μm filter.
Resazurin Solution:
Composition per 44.0mL:
Resazurin .............................................................0.011g
Preparation of Resazurin Solution:  Add resazurin to distilled/
deionized water and bring volume to 44.0mL. Mix thoroughly.
Preparation of Medium: Add components, except  L -cysteine·HCl·H2O,
to distilled/deionized water and bring volume to 100.0mL. Mix thor-
oughly. Gently heat and bring to boiling. Continue boiling until resazu-
rin turns colorless, indicating reduction. Cool in an ice-water bath
under 100% CO2 . Add the  L -cysteine·HCl·H2O. Adjust pH to 7.0 with
8N NaOH or 5N HCl. Anaerobically distribute into tubes under O2 -free
100% N2 . Cap tubes with butyl rubber stoppers. Place tubes in a press.
Autoclave for 12 min at 15 psi pressure–121°C with fast exhaust.
Use:  For the cultivation and maintenance of the Fibrobacter species.