# 1013: HALOBACTERIOVORAX MEDIUM ## Main sol. 1013 700 ml **Artificial sea water** 10 ml **Modified Wolin's mineral solution** 300 ml Distilled water 1. Mix ingredients and adjust pH of the final solution to 7.5 - 8.0. Sterilize by autoclaving at 121°C for 20 min. Grow prey bacterium (DSM 15416) on agar plates using medium 514. For 50 ml prey medium suspension use one agar plate incubated for 24 hours and wash off grown cells in 3 - 5 ml medium. The prey medium suspension should contain approx. 107 to 109 cells. Inoculate the complete medium immediately after preparation and incubate the suspension with shaking until a decrease in turbidity is visible. ## Artificial sea water 27.7 g NaCl 7 g MgSO4 x 7 H2O 5.5 g MgCl2 x 6 H2O 0.65 g KCl 0.1 g NaBr 30 mg H3BO3 15 mg SrCl2 x 6 H2O 10 mg Citric acid 0.05 mg KI 2.25 g CaCl2 x 2 H2O 1000 ml Distilled water ## 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. ## Main sol. 514 5 g Bacto peptone 1 g Yeast extract (Bacto ) 0.1 g Fe(III) citrate 19.45 g NaCl 5.9 g MgCl2 (anhydrous) 3.24 g Na2SO4 1.8 g CaCl2 0.55 g KCl 0.16 g NaHCO3 0.08 g KBr 34 mg SrCl2 22 mg H3BO3 4 mg Na-silicate 2.4 mg NaF 1.6 mg (NH4)NO3 8 mg Na2HPO4 1000 ml Distilled water 1. Final pH should be 7.6 ± 0.2 at 25°C. If using the complete medium from Difco add 37.40 g to 1 litre water.