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TD3 Medium (DSMZ Medium 876)

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

TD3 Medium

(DSMZ Medium 876)

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

Composition per 1078.0mL:

NaCl............................................................................................ 20.0g

MgCl2·6H2O ................................................................................. 9.8g

Na2SO4.......................................................................................... 4.0g

KCl................................................................................................ 0.5g

NH4Cl ......................................................................................... 0.25g

KH2PO4......................................................................................... 0.2g

CaCl2·2H2O .................................................................................. 0.1g

Resazurin ................................................................................... 0.5mg

NaHCO3 solution .....................................................................50.0mL

Na2S·9H2O solution.................................................................13.0mL

Sodium caproate solution ........................................................10.0mL

Selenite-tungstate solution.........................................................2.0mL

Seven vitamin solution ..............................................................1.0mL

Vitamin solution.........................................................................1.0mL

Trace elements solution SL-10 ..................................................1.0mL

pH 6.8 ± 0.2 at 25°C

Na2S·9H2O Solution:

Composition per 20.0mL:

Na2S·9H2O.................................................................................... 0.6g

Preparation of Na2S·9H2O Solution: Add Na2S·9H2O to dis

tilled/deionized water and bring volume to 20.0mL. Mix thoroughly.

Autoclave under 100% N2 for 15 min at 15 psi pressure–121°C. Cool

to room temperature.

NaHCO3 Solution:

Composition per 10.0mL:

NaHCO3........................................................................................ 5.0g

Preparation of NaHCO3 Solution: Add NaHCO3 to distilled/de

ionized water and bring volume to 10.0mL. Mix thoroughly. Sparge

with 80% N2 + 20% CO2. Filter sterilize.

Sodium Caproate Solution:

Composition per 10.0mL:

Sodium caproate ........................................................................... 0.5g

Preparation of Sodium Caproate Solution: Add sodium cap

roate to distilled/deionized water and bring volume to 10.0mL. Mix

thoroughly. Sparge with 100% N2. Filter sterilize.

Seven Vitamin Solution:

Composition per liter:

Pyridoxine hydrochloride ......................................................300.0mg

Thiamine-HCl·2H2O..............................................................200.0mg

Nicotinic acid.........................................................................200.0mg

Vitamin B12 ............................................................................100.0mg

Calcium pantothenate ............................................................100.0mg

p-Aminobenzoic acid...............................................................80.0mg

D(+)-Biotin...............................................................................20.0mg

Preparation of Seven Vitamin Solution: Add components to dis

tilled/deionized water and bring volume to 1.0L. Sparge with 100% N2.

Mix thoroughly. Filter sterilize.

Vitamin Solution:

Composition per 100.0mL:

Pyridoxine-HCl........................................................................10.0mg

Thiamine-HCl·2H2O..................................................................5.0mg

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

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

D-Ca-pantothenate......................................................................5.0mg

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

Lipoic acid .................................................................................5.0mg

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

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

Vitamin B12 ................................................................................0.1mg

Preparation of Vitamin Solution: Add components to distilled/

deionized water and bring volume to 100.0mL. Mix thoroughly. Sparge

with 80% H2 + 20% CO2. Filter sterilize.

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. Sparge with 80% N2 + 20% CO2. Autoclave for 15 min at 15

psi pressure–121°C.

Selenite-Tungstate Solution

Composition per liter:

NaOH ............................................................................................ 0.5g

Na2WO4·2H2O ...........................................................................4.0mg

Na2SeO3·5H2O...........................................................................3.0mg

Preparation of Selenite-Tungstate Solution: Add components

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

Sparge with 100% N2. Filter sterilize.

Optional Supplemental Fatty Acid Mixture:

Composition per 20.0mL:

Valeric acid ................................................................................... 0.5g

Isovaleric acid............................................................................... 0.5g

Alpha-Methylbutyric acid............................................................. 0.5g

Isobutyric acid .............................................................................. 0.5g

Preparation of Optional Supplemental Fatty Acid Mixture:

Add components to 20.0mL distilled/deionized water. Mix thoroughly.

Sparge with 100% N2. Filter sterilize.

Preparation of Medium: Prepare and dispense medium under 80%

N2 + 20% CO2 gas atmosphere. Add components, except NaHCO3 solu

tion, sodium caproate solution, Na2S·9H2O solution, vitamin solution,

seven vitamin solution, selenite-tungstate solution, and trace elements

solution SL-10, to distilled/deionized water and bring volume to 1.0L.

Mix thoroughly. Adjust pH to 7.2. Sparge with 80% N2 + 20% CO2. Au

toclave for 15 min at 15 psi pressure–121°C. Aseptically and anaerobi

cally add 50.0mL NaHCO3 solution, 10.0mL sodium caproate solution,

10.0mL Na2S·9H2O solution, 1.0mL vitamin solution, 1.0mL seven vi

tamin solution, 2.0mL selenite-tungstate solution, and 1.0mL trace ele

ments solution SL-10. Mix thoroughly. Aseptically and anaerobically

distribute into sterile tubes or bottles. Growth can be stimulated by addi

tion of clarified rumen fluid or 10–20mL of a mixture of fatty acids. Final

pH should be 6.7–6.9.

Use: For the cultivation of unclassified bacterium DSM 13418.

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TD3 Medium (DSMZ Medium 876)