① Put 150ml of Ringer for freshwater fish in a 300 ml beaker and put stirrer stirrer.
② Pour agar (Agar not for electrophoresis) to a concentration of 0.8-1%.
③ Cover with a saran wrap, put it on a stirrer, and stir the agar powder.
④Put the Ringer's gel in a microwave oven and warm it up so that it does not boil suddenly.
⑤ Once warmed, place in a stirrer and stir the agar flour.
⑥ Repeat ③-⑤ to dissolve the agar, keeping the solution in it from boiling. When it becomes clear, put it over the light and check that no agar powder remains.
⑦ When the agar powder is completely dissolved, cool the agar solution a little.
⑧ During this time, the glass Petri dishes are placed on a desk (checking that they are level with a level gauge).
⑨ Pour the agar liquid into a glass petri dish. Thinner for embryo manipulation, but thicker agar for photographic purposes.
⑩ When the agar solidifies, cover, invert, and store in the refrigerator.。
(9) Culture media and agarose Petri dishes
① Ringer's solution for freshwater fish: 7. 5g NaCl, 0. 2g KCl, 0. 4g CaCl2/1 l DW
② Egg White: Take the egg white from the chicken egg, make a merenge, and store it in a refrigerator. Allow to stand for a while and use the egg white fluid accumulated in the lower part. Necessary reconciliation.
③ Primary culture: Ringer's solution for freshwater fish containing 1.6% egg white
④ Primary culture solution for 96-well culture dishes: Add 1/1000 volume of antibiotic to the solution in Ringer's solution for freshwater fish containing 1.6% egg white (antibiotic solution stock: penicillin, streptomycin are dissolved in DW at 10% concentration each, and dispense 100-200 μl into test tubes to keep them frozen)
⑤ Secondary culture solution: 1. 8 mM CaCl2, 1. 8 mM MgCl2 (180 μl of 1M CaCl2, and 180 μl of 1M MgCl2 are placed in 100ml of distilled water)