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Thermoanaerobacter ethanolicus Medium

[所属分类:培养基配方] [发布时间:2021-9-6] [发布人:网站管理员2] [阅读次数:] [返回]

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

Composition per liter:

Glucose ......................................................................................... 8.0g

Na2HPO4·12H2O........................................................................... 4.2g

Yeast extract.................................................................................. 2.0g

KH2PO4......................................................................................... 1.5g

NH4Cl ........................................................................................... 0.5g

MgCl2·6H2O ............................................................................... 0.18g

Reducing solution ....................................................................40.0mL

Wolfe’s modified mineral solution ............................................5.0mL

Resazurin (0.1% solution) .........................................................1.0mL

Vitamin solution.........................................................................0.5mL

Caution: This medium contains Na2S, and H2S production will occur,

especially upon prolonged boiling. H2S is hazardous and preparation of

this medium should be done in a chemical fume hood.

Reducing Solution:

Composition per 200.0mL:

Cysteine·HCl·H2O ........................................................................ 2.5g

Na2S·9H2O.................................................................................... 2.5g

NaOH (0.2N solution)............................................................200.0mL

Preparation of Reducing Solution: Gently heat the NaOH solu

tion and bring to boiling. Gas with 95% N2 + 5% H2. Cool to room tem

perature. Add the cysteine·HCl·H2O and Na2S·9H2O. Anaerobically

distribute into tubes. Cap with rubber stoppers. Autoclave for 15 min

at 15 psi pressure–121°C.

Vitamin Solution:

Composition per 500.0mL:

Pyridoxine·HCl ............................................................................. 0.1g

p-Aminobenzoic acid.................................................................. 0.05g

Calcium pantothenate ................................................................. 0.05g

Nicotinic acid.............................................................................. 0.05g

Thioctic acid ............................................................................... 0.05g

Biotin .......................................................................................... 0.02g

Folic acid .................................................................................... 0.02g

Riboflavin .................................................................................. 5.0mg

Thiamine·HCl ............................................................................5.0mg

Vitamin B12 ................................................................................1.0mg

Preparation of Vitamin Solution: Add components to distilled/

deionized water and bring volume to 500.0mL. Mix thoroughly. Store

solution in the dark at −10°C.

Wolfe’s Modified Mineral Solution:

Composition per liter:

MgSO4·7H2O ................................................................................ 3.0g

Nitrilotriacetic acid ....................................................................... 1.5g

NaCl .............................................................................................. 1.0g

MnSO4·H2O .................................................................................. 0.5g

CaCl2 (anhydrous)......................................................................... 0.1g

Co(NO3)2·6H2O ............................................................................ 0.1g

FeSO4·7H2O.................................................................................. 0.1g

ZnSO4·7H2O ................................................................................. 0.1g

AlK(SO4)2 (anhydrous)............................................................... 0.01g

CuSO4·5H2O ............................................................................... 0.01g

H3BO3 ......................................................................................... 0.01g

Na2MoO4·2H2O .......................................................................... 0.01g

Na2SeO3 (anhydrous).................................................................1.0mg

Preparation of Wolfe’s Modified Mineral Solution: Add

nitrilotriacetic acid to 500.0mL of distilled/deionized water. Dissolve

by adjusting pH to 6.5 with KOH. Add remaining components. Add

distilled/deionized water to 1.0L.

Preparation of Medium: Add components, except reducing solu

tion, to distilled/deionized water and bring volume to 1.0L. Gently heat

and bring to boiling under 95% N2 + 5% H2. Continue boiling until col

or changes from blue to pink. Add the reducing solution. The pink col

or will disappear, indicating that the solution has been reduced.

Distribute into tubes or flasks under 95% N2 + 5% H2 using anerobic

techniques. Cap tubes with rubber stoppers. Autoclave for 15 min at 15

psi pressure–121°C.

Use: For the cultivation and maintenance of thermophilic anaerobes

such as Thermoanaerobacter species and some Clostridium species.

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Thermoanaerobacter ethanolicus Medium