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Thermodesulfotobacterium Agar

[所属分类:培养基配方] [发布时间:2021-9-7] [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:

Na2SO4........................................................................................ 30.0g

Agar ............................................................................................ 20.0g

Sodium lactate............................................................................... 4.0g

Yeast extract.................................................................................. 1.0g

Mineral solution 2....................................................................50.0mL

Na2CO3 solution......................................................................50.0mL

Mineral solution 1....................................................................25.0mL

Cysteine-sulfide reducing agent ..............................................20.0mL

Wolfe’s mineral solution..........................................................10.0mL

Wolfe’s vitamin solution..........................................................10.0mL

Resazurin (0.025% solution) .....................................................4.0mL

pH 7.2 ± 0.2 at 25°C

Mineral Solution 1:

Composition per liter:

K2HPO4......................................................................................... 6.0g

Preparation of Mineral Solution 1: Add K2HPO4 to distilled/de

ionized water and bring volume to 1.0L. Mix thoroughly.

Mineral Solution 2:

Composition per liter:

NaCl............................................................................................ 12.0g

KH2PO4......................................................................................... 6.0g

(NH4)2SO4 .................................................................................... 6.0g

MgSO4·7H2O................................................................................ 2.4g

CaCl2·2H2O .................................................................................. 1.6g

Preparation of Mineral Solution 2: Add components to distilled/

deionized water and bring volume to 1.0L. Mix thoroughly.

Na2CO3 Solution:

Composition per 100.0mL:

Na2CO3 ......................................................................................... 8.0g

Preparation of Na2CO3 Solution: Add Na2CO3 to distilled/deion

ized water and bring volume to 100.0mL. Mix thoroughly.

Cysteine-Sulfide Reducing Agent:

Composition per 20.0mL:

L-Cysteine·HCl·H2O .............................................................. 300.0mg

Na2S·9H2O............................................................................. 300.0mg

Preparation of Cysteine-Sulfide Reducing Agent: Add L

cysteine·HCl·H2O to 10.0mL of distilled/deionized water. Mix thor

oughly. In a separate tube, add Na2S·9H2O to 10.0mL of distilled/de

ionized water. Mix thoroughly. Gas both solutions with 100% N2 and

cap tubes. Autoclave both solutions for 15 min at 15 psi pressure–

121°C using fast exhaust. Cool to 50°C. Aseptically combine the two

solutions under 100% N2.

Wolfe’s Mineral Solution:

Composition per liter:

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

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

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

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

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

CoCl2·6H2O .................................................................................. 0.1g

CaCl2............................................................................................. 0.1g

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

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

AlK(SO4)2·12H2O....................................................................... 0.01g

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

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

Preparation of Wolfe’s 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.

Wolfe’s Vitamin Solution:

Composition per liter:

Pyridoxine·HCl ........................................................................10.0mg

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

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

Nicotinic acid.............................................................................5.0mg

Calcium pantothenate ................................................................5.0mg

p-Aminobenzoic acid.................................................................5.0mg

Thioctic acid ..............................................................................5.0mg

Biotin .........................................................................................2.0mg

Folic acid....................................................................................2.0mg

Cyanocobalamin .....................................................................100.0μg

Preparation of Wolfe’s Vitamin Solution: Add components to

distilled/deionized water and bring volume to 1.0L. Mix thoroughly.

Filter sterilize.

Preparation of Medium: Add components, except vitamin solu

tion and cysteine-sulfide reducing agent, to distilled/deionized water

and bring volume to 970.0mL. Mix thoroughly. Gently heat and bring

to boiling. Autoclave for 15 min at 15 psi pressure–121°C. Cool to

50°–55°C under 80% N2 + 20% CO2. Aseptically add the sterile vita

min solution and then the sterile cysteine-sulfide reducing agent. Ad

just the pH to 7.2. Distribute aseptically and anaerobically into sterile

tubes.

Use: For the cultivation and maintenance of Thermodesulfobacterium

commune and other Thermodesulfobacterium species.

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Thermodesulfotobacterium Agar