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采用PCR方法以正常中国人脐带血提取总DNA为模板，扩增出15kb的粒细胞集落刺激因子（G－CSF）基因组基因。序列分析证实其正确性。将其插入小鼠乳清酸蛋白（WAP）基因的起始密码子ATG前的Kpnl位点，使其受控于2．6kb的WAP调控序列，从而构建成乳腺表达载体pWGG。回收经EcoRI酶切后的8．7kb片段用于显微注射。共注射1200枚受精卵，移植至受体34母鼠，产生仔鼠85只。经PCR检测和Southern杂交分析，证实获得两只整合有人G－CSF基因的雄性鼠，整合率为2．37％。采用ELASA方法检测F1代雌鼠乳汁，结果表明成功地表达出人G－CSF。表达量为120～250ng／ml。这一结果表明转基因的表达具有乳腺特异性。这为在大动物中实施转基因提供了依据。
PCR method was used to amplify the 15kb G-CSF genomic DNA from the total DNA extracted from normal Chinese cord blood. Sequence analysis confirmed its correctness. Inserted into the Kpnl site before the start codon ATG of the mouse whey protein (WAP) gene and controlled by the 2.6kb WAP regulatory sequence to construct the breast expression vector pWGG. The 8.7 kb fragment digested with EcoRI was recovered for microinjection. A total of 1,200 fertilized eggs were injected into recipient 34 mothers to produce 85 offspring. PCR and Southern blot analysis confirmed that two male mice with integrated human G-CSF gene were obtained, with an overall integration rate of 2.37%. The ELASA method was used to detect the milk of F1 generation female mice, and the results showed that human G-CSF was successfully expressed. The expression level is 120 ~ 250ng / ml. This result indicates that the expression of the transgene is breast specific. This provides a basis for the implementation of GMOs in large animals.