# 1577: SPOROHALOBACTER SALINUS MEDIUM ## Main sol. 1577 1 g NH4Cl 0.5 g K2HPO4 0.1 g CaCl2 x 2 H2O 0.8 g KCl 0.5 g Na-acetate 200 g NaCl 0.5 g Yeast extract (OXOID) 5 g Tryptone (BD Bacto) 10 ml **Modified Wolin's mineral solution** 0.5 ml Sodium resazurin (0.1% w/v) 0.5 g L-Cysteine HCl x H2O 5 g MgSO4 1 g Na2CO3 4 g D-Glucose 0.5 g Na2S x 9 H2O 1000 ml Distilled water 1. Dissolve ingredients (except cysteine, magnesium sulfate, carbonate, glucose and sulfide), then sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Add and dissolve cysteine, adjust pH to 6.0, dispense under 80% N2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After sterilization add magnesium sulfate, glucose 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 gas atmosphere. Prior to inoculation check pH of complete medium and adjust to 6.8 - 7.0, if necessary. 2. Note: This medium cannot be stored over a longer period and should be prepared freshly before use! ## Modified Wolin's mineral solution 1.5 g Nitrilotriacetic acid 3 g MgSO4 x 7 H2O 0.5 g MnSO4 x H2O 1 g NaCl 0.1 g FeSO4 x 7 H2O 0.18 g CoSO4 x 7 H2O 0.1 g CaCl2 x 2 H2O 0.18 g ZnSO4 x 7 H2O 0.01 g CuSO4 x 5 H2O 0.02 g AlK(SO4)2 x 12 H2O 0.01 g H3BO3 0.01 g Na2MoO4 x 2 H2O 0.03 g NiCl2 x 6 H2O 0.3 mg Na2SeO3 x 5 H2O 0.4 mg Na2WO4 x 2 H2O 1000 ml Distilled water 1. First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH.