323. PSYCHROPHYLIC CLOSTRIDIUM MEDIUM

Tryptone 10.0 g

Gelatin peptone 10.0 g

Yeast extract 5.0 g

D-glucose 1.0 g

Sodium chloride 5.0 g

L-Arginine 1.0 g

Sodium pyruvate 1.0 g

Menadione 0.5 mg

Haemin 5.0 mg

Distilled water 990 ml

Dissolve all components in distilled water and add 0.5 ml/l Na-resazurin solution (0.1% w/v), bring to the boil and cool to room temperature while sparging with 80% N₂ and 20% CO₂ gas mixture. Add L-cysteine-HCl × H₂O (0.3 g/l), dispense the medium into anoxic Hungate-type tubes or serum vials and autoclave. After autoclaving supplement medium with 5.0 g/l D-glucose added from a sterile anoxic stock solution sterilized by filtration. Adjust pH of medium to 6.8 with a sterile anoxic stock solution of Na₂CO₃ (5% w/v) prepared under 80% N₂ and 20% CO₂ gas atmosphere.

 

Updated 01.02.2024