Haloanaerobium salsugo Medium

[所属分类:培养基配方] [发布时间:2021-3-24] [发布人: ] [阅读次数:] [返回]

Haloanaerobium salsugo Medium

培养基配方

Composition per liter:
NaCl............................................................................................90.0g
Purified agar (if necessary).........................................................20.0g

Casamino acids.............................................................................5.0g

Yeast extract..................................................................................5.0g
Dipotassium PIPES (piperazine-N,N′-
bis[2-ethanesulfonic acid]) buffer ..........................................1.5g
Resazurin ...................................................................................1.0mg
Glucose solution ......................................................................50.0mL
L -Cysteine-sulfide reducing solution.......................................20.0mL
Mineral solution.......................................................................20.0mL
Wolfe’s vitamin solution..........................................................10.0mL
Modified Wolfe’s mineral solution............................................5.0mL
pH 6.0–7.0 at 25°C
Glucose Solution:
Composition per 50.0mL:
D -Glucose....................................................................................10.0g
Preparation of Glucose Solution:  Add glucose to distilled/deionized water and bring volume to 50.0mL. Mix thoroughly. Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C.
Mineral Solution:
Composition per liter:
NH 4 Cl .........................................................................................50.0g
NaCl............................................................................................40.0g
MgSO 4 ·7H 2 O..............................................................................10.0g
KCl................................................................................................5.0g
KH 2 PO 4 .........................................................................................5.0g
CaCl 2 ·2H 2 O.................................................................................. 2.0g
Preparation of Mineral Solution:  Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly.
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.
Modified Wolfe’s Mineral Solution:
Composition per liter:
MgSO 4 ·7H 2 O................................................................................3.0g
Nitrilotriacetic acid.......................................................................1.5g
NaCl..............................................................................................1.0g
MnSO 4 ·H 2 O..................................................................................0.5g
CaCl 2 .............................................................................................0.1g
CoCl 2 ·6H 2 O ..................................................................................0.1g
FeSO 4 ·7H 2 O..................................................................................0.1g
ZnSO 4 ·7H 2 O.................................................................................0.1g
AlK(SO 4 ) 2 ·12H 2 O.......................................................................0.01g
CuSO 4 ·5H 2 O...............................................................................0.01g
H 3 BO 3 .........................................................................................0.01g
Na 2 MoO 4 ·2H 2 O ..........................................................................0.01g
Na 2 SeO 3 ......................................................................................0.01g
NaWO 4 ·2H 2 O..............................................................................0.01g
NiC1 2 ·6H 2 O................................................................................0.01g
Preparation of Modified Wolfe’s Mineral Solution: Add nitrilotriacetic acid to 500.0mL of distilled/deionized water. Adjust pH to 6.5 with KOH. Add remaining components one at a time. Add distilled/deionized water to 1.0L. Adjust pH to 6.8.
L -Cysteine-Sulfide Reducing Solution:
Composition per 200.0mL:
L -Cysteine·HCl·H 2 O.....................................................................5.0g
Na 2 S·9H 2 O....................................................................................5.0g
NaOH..........................................................................................1.25g
Preparation of  L -Cysteine-Sulfide Reducing Solution: Add NaOH to distilled/deionized water and bring volume to 200.0mL. Mix thoroughly. Gently heat and bring to boiling. Cool to room temperature while sparging with 100% N 2 . Add  L -cysteine·HCl·H 2 O and Na 2 S·9H 2 O. Mix thoroughly. Anaerobically distribute into tubes. Autoclave for 15 min at 15 psi pressure–121°C.
Preparation of Medium: Prepare and dispense medium under 100% N 2 . Add components, except glucose solution and  L -cysteinesulfide reducing solution, to distilled/deionized water and bring volume to 950.0mL. Mix thoroughly. Gently heat and bring to boiling. Continue boiling for 3 min. Cool to room temperature while sparging with 100% N 2 . Adjust pH to 6.0–7.0. Add 20.0mL of  L -cysteine-sulfide reducing solution. Mix thoroughly. Anaerobically distribute 9.5mL volumes into anaerobic tubes. Autoclave for 15 min at 15 psi pressure–121°C. Aseptically and anaerobically add 0.5mL of sterile glucose solution to each tube. Mix thoroughly.
Use: For the cultivation of Haloanaerobium salsugo.