# 383a: DESULFOTIGNUM MEDIUM ## Main sol. 383a 962 ml **Solution A** 20 ml **Solution B** 4 ml **Solution C** 1 ml **Solution D** 13 ml **Solution E** 1. Solution A is sparged with 80% N2 and 20% CO2 gas mixture to reach a pH below 6 (at least 30 min), then dispensed under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclaved. Solution B is autoclaved separately under 80% N2 and 20% CO2 gas atmosphere. Solutions C and D are prepared under 100% N2 gas and filter-sterilized. Solution E is autoclaved under 100% N2 gas. To complete the medium appropriate amounts of solutions B to E are added to the sterile solution A in the sequence as indicated. Final pH of the medium should be 7.0 – 7.2. 2. Note: Addition of 10 - 20 mg sodium dithionite per liter (e.g. from 5% (w/v) solution freshly prepared under N2 and filter-sterilized) may stimulate growth of some strains at the beginning. For transfers use 5 - 10% inoculum. Incubate all strains in the dark. ## Solution A 3 g Na2SO4 0.2 g KH2PO4 0.3 g NH4Cl 15 g NaCl 3 g MgCl2 x 6 H2O 0.5 g KCl 0.15 g CaCl2 x 2 H2O 1 ml **Trace element solution SL-10** 1 ml **Selenite-tungstate solution** 0.5 ml Sodium resazurin (0.1% w/v) 960 ml Distilled water ## Solution B 1 g Na2CO3 20 ml Distilled water ## Solution C 1 g Na-pyruvate 4 ml Distilled water ## Solution D 1 ml **Wolin's vitamin solution (10x)** ## Solution E 0.4 g Na2S x 9 H2O 13 ml Distilled water ## Selenite-tungstate solution 0.5 g NaOH 3 mg Na2SeO3 x 5 H2O 4 mg Na2WO4 x 2 H2O 1000 ml Distilled water ## 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. ## 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