# 1210: GEOGLOBUS MEDIUM ## Main sol. 1210 18 g NaCl 4 g MgCl2 x 6 H2O 0.33 g KCl 0.33 g CaCl2 x 2 H2O 0.5 g (NH4)2SO4 1 ml **Trace element solution SL-10** 1 ml **Selenite-tungstate solution** 200 ml **Amorphous Fe(OH)3** 0.5 ml Sodium resazurin (0.1% w/v) 1 g Na2CO3 1.5 g Na-acetate 0.33 g KH2PO4 0.2 g Yeast extract 1 ml **Wolin's vitamin solution (10x)** 800 ml Distilled water 1. Dissolve ingredients except carbonate, acetate, hydrogenphosphate, yeast extract, vitamins and ferric iron hydroxide sludge. Suspend pellet of ferric iron hydroxide in medium and sparge with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Thereafter, dispense suspension under 80% N2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave for 30 min. Add acetate, hydrogenphosphate, yeast extract and vitamins (sterilized by filtration) 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 mixture. The pH of the complete medium should be at 6.5 - 6.8. ## Trace element solution SL-10 10 ml HCl (25%) 1.5 g FeCl2 x 4 H2O 70 mg ZnCl2 100 mg MnCl2 x 4 H2O 6 mg H3BO3 190 mg CoCl2 x 6 H2O 2 mg CuCl2 x 2 H2O 24 mg NiCl2 x 6 H2O 36 mg Na2MoO4 x 2 H2O 990 ml Distilled water 1. First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml. ## Selenite-tungstate solution 0.5 g NaOH 3 mg Na2SeO3 x 5 H2O 4 mg Na2WO4 x 2 H2O 1000 ml Distilled water ## Amorphous Fe(OH)3 320 ml FeCl3 x 6 H2O (60.00 g/l) 80 ml NaOH (10% w/v), adjust if required 1. Slowly titrate 320 ml of a FeCl3 x 6 H2O stock solution (60.00 g/l) with 10% (w/v) NaOH to pH 8.0-8.5 under agitation (use magnetic stirrer). Total amount of added NaOH approx. 80 - 100 ml. The precipitated Fe(OH)3 should be stored at room temperature overnight with surface covered with water. Thereafter, centrifuge at 2000 rpm for 5 min and discard the supernatant. Wash several times with distilled water. Resuspend the pellet in medium as described above. For storage autoclave under 100% N2 atmosphere. ## Wolin's vitamin solution (10x) 20 mg Biotin 20 mg Folic acid 100 mg Pyridoxine hydrochloride 50 mg Thiamine HCl 50 mg Riboflavin 50 mg Nicotinic acid 50 mg Calcium D-(+)-pantothenate 1 mg Vitamin B12 50 mg p-Aminobenzoic acid 50 mg (DL)-alpha-Lipoic acid 1000 ml Distilled water