CAJ | 학술논문

[目的]旨在探讨通过向水牛受精卵胞质内注射外源DNA实现转基因的可行性。[方法]水牛卵母细胞体外成熟20～22h后随机分为2组,①在体外受精7～10或18～20h后向卵胞质内注入约7.5pL50μg/ml含线性EGFP片段的DNA溶液;②则分别注入单个精子与约7.5pL50μg/ml含线性EGFP片段的DNA混合物,观察外源基因在胚胎发育过程中的表达情况。[结果]受精卵胞质内注射的早期胚胎基因表达率、囊胚基因表达率与ICSI-Tr差异不显著（P〉0.05）,且IVF7～10h时注射的分裂率、早期胚胎基因表达率均显著高于18～20h（P〈0.05）。[结论]水牛IVF受精卵胞质内注射外源基因能获得转基因胚胎,且IVF后7～10h注射的效果优于IVF后18～20h注射。
[목적]지재탐토통과향수우수정란포질내주사외원DNA실현전기인적가행성。[방법]수우란모세포체외성숙20～22h후수궤분위2조,①재체외수정7～10혹18～20h후향란포질내주입약7.5pL50μg/ml함선성EGFP편단적DNA용액;②칙분별주입단개정자여약7.5pL50μg/ml함선성EGFP편단적DNA혼합물,관찰외원기인재배태발육과정중적표체정황。[결과]수정란포질내주사적조기배태기인표체솔、낭배기인표체솔여ICSI-Tr차이불현저（P〉0.05）,차IVF7～10h시주사적분렬솔、조기배태기인표체솔균현저고우18～20h（P〈0.05）。[결론]수우IVF수정란포질내주사외원기인능획득전기인배태,차IVF후7～10h주사적효과우우IVF후18～20h주사。
[Objective] This study aimed to investigate the feasibility of transgenesis by injecting exogenous DNA into zygote cytoplasm of Buffalo. [Method] Buffalo oocytes were randomly divided into two groups 20-22 h after in vitro maturation. One group of oocytes was introduced with about 7.5 pl of 50 μg/ml DNA solution containing linear EGFP fragment by cytoplasmic injection 7-10 h or 18-20 h after in vitro fertilization （IVF）; the other group of oocytes was introduced with mixture of a single buffalo sperm and about 7.5 pl of 50 μg/ml DNA solution containing linear EGFP fragment by cytoplasmic injection （generally called ICSI-Mediated Gene Transfer, ICSI-Tr）. Expression of exogenous DNA was observed and recorded during the process of embryonic development. [Result] Early embryonic gene expression efficiency and blastocyst gene expression efficiency in IVF injection group showed no significant difference compared with that in ICSI-Tr group （P0.05）. In addition, the cleavage rate and early embryonic gene expression efficiency in IVF injection group were significantly higher with injection at 7-10 h post IVF than that at 18-20 h post IVF （P0.05）. [Conclusion] These results indicate that transgenic buffalo embryos can be generated by injecting exogenous DNA into cytoplasm of IVF oocytes, and the optimal injection time is 7-10 h post IVF.