Clostridium halophilum Medium

[所属分类:培养基配方] [发布时间:2020-5-11] [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

Clostridium halophilum Medium

培养基配方
Composition per liter:
Solution A....................................................................900.0mL
Solution B.....................................................................80.0mL
Solution C.....................................................................10.0mL
Solution D.....................................................................10.0mL
pH 8.3 ± 0.2 at 25°C
Solution A:
Composition per 900.0mL:
NaCl............................................................................60.0g
Betaine.........................................................................5.86g
MgSO4 ·7H2O.....................................................................5.0g
L -Alanine ......................................................................2.2g
NH4Cl ...........................................................................1.0g
Yeast extract....................................................................1.0g
CaC12 ·2H2O...................................................................25.0mg
Resazurin ......................................................................1.0mg
Na2SeO3 ·5H2O................................................................15.0μg
Wolfe’s vitamin solution......................................................10.0mL
Trace elements solution SL-10...................................................1.0mL
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-10:
Composition per liter:
FeCl2 ·4H2O.....................................................................1.5g
CoCl2 ·6H2O..................................................................190.0mg
MnCl2 ·4H2O .................................................................100.0mg
ZnCl2 .........................................................................70.0mg
Na2MoO4 ·2H2O.................................................................36.0mg
NiCl2 ·6H2O...................................................................24.0mg
H3BO3 ..........................................................................6.0mg
CuCl2 ·2H2O....................................................................2.0mg
HCl (25% solution).............................................................10.0mL
Preparation of Trace Elements Solution SL-10: 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 Solution A: Prepare and dispense medium under
80% N2 + 20% CO2 . Add components to distilled/deionized water and
bring volume to 900.0mL. Mix thoroughly. Gently heat and bring to
boiling. Continue boiling for 3 min. Cool to room temperature while
sparging with 80% N2 + 20% CO2 .
Solution B:
Composition per 80.0mL:
NaHCO3 ........................................................................5.0g
Preparation of Solution B: Add NaHCO3 to distilled/deionized
water and bring volume to 80.0mL. Mix thoroughly. Sparge with 100%
N2 for 20 min.
Solution C:
Composition per 10.0mL:
K2HPO4 .......................................................................0.358g
KH2PO4 .......................................................................0.223g
Preparation of Solution C: Add components to distilled/deionized
water and bring volume to 10.0mL. Mix thoroughly. Sparge with 100%
N2 . Autoclave for 15 min at 15 psi pressure–121°C.
Solution D:
Composition per 10.0mL:
Na2S·9H2O.....................................................................0.3g
Preparation of Solution D: Add components to distilled/deionized
water and bring volume to 10.0mL. Mix thoroughly. Sparge with 100%
N2 . Autoclave for 15 min at 15 psi pressure–121°C.
Preparation of Medium: Prepare and dispense medium under 80%
N2 + 20% CO2 . Anaerobically combine 900.0mL of cooled solution A
with 80.0mL of sparged solution B. Mix thoroughly. Adjust pH to 8.3.
Anaerobically distribute 9.8mL volumes into anaerobe tubes. Auto-
clave for 15 min at 15 psi pressure–121°C. Aseptically and anaerobi-
cally add 0.1mL of sterile solution C and 0.1mL of sterile solution D to
each tube. Mix thoroughly.
Use: For the cultivation of Clostridium halophilum.