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通过正压手动显微操作技术 ,将能在离体细胞和活体细胞表达的人凝血因子Ⅸ的表达载体pMCⅨmDNA导入 586枚昆明白种小鼠受精卵 ,随后将显微注射后存活的 4 99枚受精卵移植于假孕受体母鼠 ,获得2 16只F0 代仔鼠 .PCR和Southern印迹杂交分析证实有 6只仔鼠 (2♀ ,4♂ )的基因组中有外源基因阳性整合 ,整合率为 3% (6 / 2 16 ) ;ELISA测定 2只F0 代转基因雌鼠的乳汁中人凝血因子Ⅸ蛋白 (hFⅨ )的含量均已达 10 0ng/mL以上 ,一期法测定其凝血活性分别为 4 4 6 7%和 79 4 3% .
The expression vector pMCIXmRNA of human coagulation factor IX, which can be expressed in ex vivo cells and living cells, was introduced into 586 Kunming white mouse zygotes by positive pressure manual micromanipulation technique, followed by 997 The fertilized eggs were transplanted into fake pregnant female mice to obtain 2 16 F0 offspring rats.PCR and Southern blot analysis confirmed the positive integration and integration of exogenous genes in the genomes of 6 offspring (2 ♀ and 4 ♂) The rate of 3% (6/216) was determined by ELISA. The content of human coagulation factor Ⅸ protein (hFⅨ) in breast milk of two F0 transgenic mice was above 10 0 ng / mL by ELISA. 4 4 6 7% and 79 4 3%.