マイクロインジェクション法による外来遺伝子導入に適したメダカ卵母細胞の成熟段階〔英文〕

概要

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Microinjection of foreign DNA into oocyte nucleus is a useful method for producing transgenic medaka. To find the most appropriate stage of oocyte maturity to obtain efficient foreign gene transfer, we microinjected the plasmid pHV-GH containing Drosophila Hsp70 promoter and rainbow trout growth hormone cDNA into medaka oocytes collected 20-6h before ovulation at intervals of 2 hours. Injected oocytes were cultured until ovulation, inseminated and incubated for 7 days. Oocytes collected 14-6h before ovulation matured in culture while most of those col-lected 20-16h before ovulation did not. The proportion of oocytes which were successfully fertilized and normally developed to the 7-day-old stage after microinjection were highest for oocytes collected at 8h before ovulation, as 79% were successfully fertilized and 58% developed normally. Embryos developed normally to the 7-day-old stage were assayed for the existence of foreign sequences by slot blotting. The proportion of embryos positive for foreign sequences among those assayed was highest (60%) for oocytes at 8 and 10h before ovulation. Thus, the gene transfer efficiency, ie. the ratio of positive embryos to injected oocytes, was highest (35%) at 8h before ovulation.
日本水産學會の論文