# 144b: CALDICELLULOSIRUPTOR OWENSIS MEDIUM ## Main sol. 144b 0.9 g NH4Cl 0.9 g NaCl 0.1 g MgCl2 x 6 H2O 0.05 g CaCl2 x 2 H2O 0.4 g K2HPO4 10 ml **Trace element solution** 3 g Yeast extract 10 g Trypticase peptone (BD BBL) 0.5 ml Sodium resazurin (0.1% w/v) 2 g Na2CO3 5 ml **Wolin's vitamin solution** 5 g D-Glucose 1 g Na2S x 9 H2O 1000 ml Distilled water 1. Dissolve ingredients (except carbonate, vitamins, glucose and sulfide), sparge medium with 80% N2 and 20% CO2 gas for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add glucose, vitamins (sterilized by filtration) and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2. Adjust pH of complete medium to 7.2 - 7.4. ## Trace element solution 0.5 g Na2-EDTA x 2 H2O 0.15 g CoCl2 x 6 H2O 0.1 g MnCl2 x 4 H2O 0.1 g FeSO4 x 7 H2O 0.1 g ZnCl2 40 mg AlCl3 x 6 H2O 40 mg Na2WO4 x 2 H2O 30 mg Na2SeO3 x 5 H2O 20 mg NiCl2 x 6 H2O 20 mg CuCl2 x 2 H2O 10 mg H3BO3 10 mg Na2MoO4 x 2 H2O 1000 ml Distilled water 1. First dissolve EDTA in distilled water and adjust pH to 7 using 2 N NaOH, then dissolve remaining compounds. ## Wolin's vitamin solution 2 mg Biotin 2 mg Folic acid 10 mg Pyridoxine hydrochloride 5 mg Thiamine HCl 5 mg Riboflavin 5 mg Nicotinic acid 5 mg Calcium D-(+)-pantothenate 0.1 mg Vitamin B12 5 mg p-Aminobenzoic acid 5 mg (DL)-alpha-Lipoic acid 1000 ml Distilled water