摘要
Maturation process of zebrafish oocyte was investigated using in vitro incubation.In medium EM-199 containing 0.5 μg/ml of 17α-hydroxyprogesterone incubated under 80％ O_2 and at 25°C,germinal vesicles(GV)of oocytes in stage Ⅳ migrated from midway between the center and theperiphery ofoocytes to the periphery in 40 minutes and the oocytes went into stage V.Half an hourlater,the oocytes underwent germinel vesicle breakdown(GVBD)with a breakdown rate of 59％.Two more hours were needed for such oocytes to complete their final maturation.The mature eggscould not come off from the follicle layer surrounding them by themselves(ovulation).By removingthe follicle and adding active sperms for insemination,we could make the mature eggs fertilized.Thechorion was elevated and blastoderm formed on the animal pole.The cleavage and development ofthese fertilized eggs followed the same course as the naturally matured and fertilized eggs.Usingblastula formation as a marker of successful fertilization of the in vitro matured egg,the fertilizationrate was 78％.This is the first report on the successful in vitro incubation of mature oocytes inzebrafish.The establishment of this in vitro oocyte maturation technology has laid the foundationfor further investigation of the transfer of foreign genes in the germinal vesicles of oocytes.
Maturation process of zebrafish oocyte was investigated using in vitro incubation.In medium EM- 199 containing 0.5 μg/ml of 17α-hydroxyprogesterone incubated under 80％ O_2 and at 25°C, germinal vesicles(GV)of oocytes in stage Ⅳ migrated from midway between the center and the periphery ofoocytes to the periphery in 40 minutes and the oocytes went into stage V.Half an hour later,the oocytes underwent germinel vesicle breakdown(GVBD)with a breakdown rate of 59％. Two more hours were needed for such oocytes to complete their final maturation.The mature eggs could not come off from the follicle layer surrounding them by themselves(ovulation).By removing the follicle and adding active sperms for insemination,we could make the mature eggs fertilized.The chorion was elevated and blastoderm formed on the animal pole.The cleavage and development of these fertilized eggs followed the same course as the naturally matured and fertilized eggs.Using blastula formation as a marker of successful fertilization of the in vitro matured egg,the fertilization rate was 78％.This is the first report on the successful in vitro incubation of mature oocytes in zebrafish.The establishment of this in vitro oocyte maturation technology has laid the foundation for further investigation of the transfer of foreign genes in the germinal vesicles of oocytes.
基金
1.This work was supported by grants from the Chinese National Natural Science Foundation(No.3897049),the Key Project of Biotechnology of the Chinese Academy of Sciences(No.75710506),the Chinese National High-Tech Project(No.8631010833).
