# J983: ARTIFICIAL SEAWATER MEDIUM WITH PULLULAN ## Main sol. J983 80 g NaCl 25 g MgSO4 x 7 H2O 4.5 g CaCl2 x 2 H2O 1.5 g KCl 1 ml **FeCl2 solution** 1 ml **Trace element solution** 1 ml **Selenite-tungstate solution** 0.5 mg Resazurin 0.5 g Pullulan 1000 ml Distilled water 1. Mix components thoroughly and autoclave under a N2-CO2 (4:1, v/v) gas mixture. Aseptically and anaerobically add the following solutions (autoclaved or *filter-sterilized and stored under N2): 10 ml NH4Cl (1%) 5 ml KH2PO4 (1%) 10 ml **Trace vitamins** (*) 20 ml Na2CO3 (5%) 6 ml Na2S x 9 H2O (5%) 2. Adjust pH to 7.2 - 7.5 with a 5% Na2CO3 solution, if necessary. ## FeCl2 solution 10 ml HCl (7.7 M, 25%) 1.5 g FeCl2 x 4 H2O 990 ml Distilled water ## Trace element solution 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 1000 ml Distilled water ## Selenite-tungstate solution 0.4 g NaOH 6 mg Na2SeO3 x 5 H2O 8 mg Na2WO4 x 2 H2O 1000 ml Distilled water ## Trace vitamins 2 mg Biotin 2 mg Folic acid 10 mg Pyridoxine hydrochloride 5 mg Thiamine HCl 5 mg Riboflavin 5 mg Nicotinic acid 5 mg Calcium pantothenate 0.1 mg Vitamin B12 5 mg p-Aminobenzoic acid 5 mg Lipoic acid 1000 ml Distilled water