NaCl 120.00 g K2HPO4 1.50 g NH4Cl 0.50 g KNO3 1.00 g Trace elements solution (Pfennig, 1965) 1.00 ml CaCl2 x 2 H2O 0.05 g MgSO4 x 7 H2O 0.50 g Na2S2O3 x 5 H2O 5.00 g NaHCO3 5.00 g Seven vitamins solution 1.00 ml Distilled water 1000.00 ml
1. Dissolve sodium chloride, potassium hydrogenphosphate, ammonium chloride and potassium nitrate, then sparge solution with 80% N2 and 20% CO2 gas mixture for at least 30 - 45 min to remove dissolved oxygen. Dispense solution under same gas atmosphere in vials suitable for anaerobic cultures (e.g. Balch-type tubes) to 50% volume, close vials with butyl rubber septa and autoclave. Add trace elements, calcium chloride, magnesium sulfate, thiosulfate, and vitamins from sterile anoxic stock solutions prepared under 100% N2 gas and bicarbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Thiosulfate and vitamins should be sterilized by filtration. Adjust pH of the complete medium to 7.5 - 7.8 using a sterile anoxic stock solution of sodium carbonate (5% w/v) prepared under 80% N2 and 20% CO2 gas atmosphere. 2.Note: Use at least 10% (v/v) as inoculum. For DSM 16925: Reduce amount of thiosulfate to 1.00 g/l.
Trace elements solution (Pfennig, 1965) (from medium 1369) EDTA 5.00 g FeSO4 x 7 H2O 2.20 g ZnSO4 x 7 H2O 0.10 g MnCl2 x 4 H2O 0.03 g H3BO3 0.03 g CoCl2 x 6 H2O 0.20 g CuCl2 x 2 H2O 0.03 g NiCl2 x 6 H2O 0.03 g Na2MoO4 x 2 H2O 0.03 g Distilled water 1000.00 ml pH 3.0-4.0 Seven vitamins solution (from medium 503) Vitamin B12 100.00 mg p-Aminobenzoic acid 80.00 mg D-(+)-biotin 20.00 mg Nicotinic acid 200.00 mg Calcium pantothenate 100.00 mg Pyridoxine hydrochloride 300.00 mg Thiamine-HCl x 2 H2O 200.00 mg Distilled water 1000.00 ml
