# 113: THIOBACILLUS DENITRIFICANS MEDIUM ## Main sol. 113 962 ml **Solution A** 20 ml **Solution B** 20 ml **Solution C** 1 ml **Solution D** 1. Solutions A, B and D are sterilized separately by autoclaving at 121°C for 15 min under 100% N2 gas atmosphere. Solution C is sterilized by filtration under an atmosphere of 80% N2 and 20% CO2 gas mixture. Appropriate amounts of solutions B to D are added to the sterile solution A in the sequence as indicated. 2. Note: A small amount of white precipitate forms after autoclaving, but this has no negative effect on growth. ## Solution A 2 g KH2PO4 2 g KNO3 1 g NH4Cl 0.8 g MgSO4 x 7 H2O 2 ml **Trace element solution SL-4** 15 g Agar, for solid medium (optional) 960 ml Distilled water 1. Adjust pH to 7.0 with NaOH. ## Solution B 5 g Na2S2O3 x 5 H2O 20 ml Distilled water ## Solution C 1 g NaHCO3 20 ml Distilled water ## Solution D 2 mg FeSO4 x 7 H2O 1 ml H2SO4 (0.1 N) ## Trace element solution SL-4 0.5 g Na2-EDTA 0.2 g FeSO4 x 7 H2O 0.1 g ZnSO4 x 7 H2O 0.03 g MnCl2 x 4 H2O 0.3 g H3BO3 0.2 g CoCl2 x 6 H2O 0.01 g CuCl2 x 2 H2O 0.02 g NiCl2 x 6 H2O 0.03 g Na2MoO4 x 2 H2O 1000 ml Distilled water 1. First dissolve EDTA in distilled water and adjust pH to 7.0 using 2 N NaOH; then add other compounds.