GSM8395535: znf706_stageII_oocyte_rep2; Danio rerio; RNA-Seq
In the context of the current investigation, a protocol for the isolation of oocytes from juvenile female fish was developed, aimed at maximizing yield and maintaining cellular integrity. The first step involved the euthanization of the WT and znf706-/- juvenile female fish (3-month-old) by submerging them in an ice-water mixture. After euthanization, the abdominal cavity of the fish was carefully opened using surgical scissors and the ovaries were bilaterally detached from the abdomen by utilizing forceps. The excised ovaries were promptly transferred into a 6-well plate preloaded with 2 ml digestive buffer (cell dissociation buffer, PCM, PC-33689) and incubated for 2-3 hrs at a temperature of 28.5℃. To enhance the efficacy of the digestive process, the 6-well plate was shaken every half hour during incubation. Once the digestion period was complete, the L-15 medium, which had been preheated to 28.5℃, was added to the digestive buffer to halt the digestion process effectively. To eliminate residual impurities, the oocytes were subjected to a series of washing steps involving the addition of fresh L-15 medium, followed by gentle resuspension of the oocytes, a process repeated 2-3 times to ensure thorough cleansing. Approximately 600 stage I oocytes and 150 stage II oocytes were collected manually with a pipette. All the liquid nitrogen-freezing samples (stages I and II oocytes) were sent to Novogene for RNA-seq library preparation