# C108: ZM/10 ## Mail sol. C108 0.5 g Bacto peptone 0.1 g Yeast extract 15 g Agar (Bacto) 10 ml **Marine Supplement Stock** 750 ml Sea water (filtered) 250 ml Deionized water 1. Make up to 1 litre with 75% filtered seawater: 25% deionized water. Autoclave at 15 psi for 15 minutes. Cool to approx. 55 °C and add sterile marine supplement. ## Marine Supplement Stock 1. First prepare primary stocks for marine supplement stock: 1 ml **Trace elements (x10 concentration)** 1 ml **Vitamin mix** 1 ml Na2SeO3 (0.002 g/l stock solution) 2. Autoclave at 15 psi for 15 minutes. Once at room temperature add Primary stock 2 (Vitamin mix). ## Trace elements (x10 concentration) 8.32 g Na2-EDTA 6.3 g FeCl3 x 6 H2O 0.02 g CuSO4 x 5 H2O 0.044 g ZnSO4 x 7 H2O 0.02 g CoCl2 x 6 H2O 0.36 g MnCl2 x 4 H2O 0.012 g Na2MoO4 x 2 H2O ## Vitamin mix 0.001 g Cyanocobalamin 0.02 g Biotin 0.05 g Thiamine HCl 0.05 g Calcium pantothenate 0.02 g Folic acid 0.05 g Riboflavin 0.05 g Nicotinamide 1. Dissolve vitamins one at a time; adjust the pH to 7 with 1 M NaOH or 1 M HCl if necessary, to dissolve the biotin and folic acid. Filter sterilise. Dispense excess into 1 ml aliquots and freeze.