[所屬分類:培養(yǎng)基配方] [發(fā)布時間:2020-7-9] [發(fā)布人:] [閱讀次數(shù):] [返回]

山東拓普生物工程有限公司

Shandong Tuopu Biol-Engineering Co.,Ltd

Eubacterium callanderi Medium

培養(yǎng)基配方
Composition per liter:
NaHCO3 ....................................................................3.5g
Glucose ...................................................................1.8g
Resazurin ................................................................1.0mg
Clarified rumen fluid ...................................................50.0mL
Pfennig's mineral solution...............................................50.0mL
L -Cysteine-sulfide reducing agent.......................................20.0mL
Wolfe’s vitamin solution................................................10.0mL
Trace elements solution SL-7 .............................................1.0mL
Pfennig’s Mineral Solution:
Composition per 100.0mL:
KH2PO4 ...................................................................1.0g
NaCl......................................................................0.8g
NH4Cl ....................................................................0.8g
MgCl2 ·6H2O.............................................................0.66g
CaCl2 ·2H2O..............................................................0.1g
Preparation of Pfennig’s Mineral Solution: Add components to
distilled/deionized water and bring volume to 100.0mL. Mix thorough-
ly.
L -Cysteine-Sulfide Reducing Agent:
Composition per 200.0mL:
L -Cysteine·HCl·H2O......................................................2.5g
Na2S·9H2O.................................................................2.5g
NaOH (3N solution)........................................................13.4mL
Preparation of  L -Cysteine-Sulfide Reducing Agent: Add  L -
cysteine·HCl·H2O to distilled/deionized water and bring volume to
50.0mL. Rapidly adjust pH to 10 with 3N NaOH solution. Add
Na2S·9H2O. Mix thoroughly. Bring volume to 200.0mL with distilled/
deionized water. Gently heat and bring to boiling. Cool to room tem-
perature. Distribute into tubes under 100% N2 . Autoclave for 15 min at
15 psi pressure–121°C.
Wolfe’s Vitamin Solution:
Composition per liter:
Pyridoxine·HCl.........................................................10.0mg
p-Aminobenzoic acid.....................................................5.0mg
Lipoic acid.............................................................5.0mg
Nicotinic acid..........................................................5.0mg
Riboflavin..............................................................5.0mg
Thiamine·HCl...........................................................5.0mg
Calcium  DL -pantothenate...............................................5.0mg
Biotin..................................................................2.0mg
Folic acid .............................................................2.0mg
Vitamin B 12 ...........................................................0.1mg
Preparation of Wolfe’s Vitamin Solution:  Add components to
distilled/deionized water and bring volume to 1.0L. Mix thoroughly.
Trace Elements Solution SL-7:
Composition per 1010.0mL:
FeCl2 ·4H2O...........................................................1.5g
CoCl2 ·6H2O...........................................................190.0mg
MnCl2 ·4H2O ..........................................................100.0mg
ZnCl2 .................................................................70.0mg
H3BO3 .................................................................62.0mg
Na2 MoO4 ·2H2O........................................................36.0mg
NiCl2 ·6H2O...........................................................24.0mg
CuCl2 ·2H2O...........................................................17.0mg
Hydrochloric acid, 25%.................................................10.0mL
Preparation of Trace Elements Solution SL-7: Add FeCl2 ·4H2O
to 10.0mL of HCl solution. Mix thoroughly. Add distilled/deionized
water and bring volume to 1.0L. Add remaining components. Mix thor-
oughly.
Preparation of Medium: Prepare and dispense medium under 80%
N2 + 20% CO2 . Add components, except NaHCO 3 and cysteine-sulfide
reducing agent, to distilled/deionized water and bring volume to
980.0mL. Mix thoroughly. Gently heat and bring to boiling. Continue
boiling for 3 min. Cool to room temperature while sparging with 80%
N2 + 20% CO2 . Add NaHCO3 . Mix thoroughly. Anaerobically distrib-
ute 10.0mL volumes into anaerobic tubes. Autoclave for 15 min at 15
psi pressure–121°C. Aseptically and anaerobically add 0.2mL of ster-
ile cysteine-sulfide reducing agent to each tube. Mix thoroughly.
Use: For the cultivation of Eubacterium callanderi.