膜反应性溶解抑制物在细胞及小鼠体内的表达

Expression of human CD59 protein in transgenic pig aortic endothelial cells and mice

目的观察外源基因在猪血管内皮细胞及小鼠体内表达情况。方法体外培养猪血管内皮细胞，脂质体转染法将pcDNA3-ICAM-2- EnhancerCD59cDNA转染猪的血管内皮细胞，以未转染的猪血管内皮细胞为阴性对照，人血细胞为阳性对照，经G418筛选（浓度为100mg／L）获得具有抗性的克隆，通过流式细胞仪检测转染细胞的表达情况。通过显微注射将CD59基因片段导入小鼠授精卵，对出生的小鼠进行基因检测。结果pcDNA3-ICAM-2-Enhancer-CD59cDNA重组质粒在细胞中表达强度为63．1％；转CD59基因小鼠外周血单核细胞（PBMCs）表达强度为73．38％。结论以人细胞间黏附因子Ⅱ（ICAM-2）为启动子并带增强子的CD59重组基因在猪的血管内皮细胞及小鼠体内均高效特异表达。

Objective To investigate the expression of human CD59 protein in transgenic pig aortic endothelial cells （PAECs） and mice. Methods The PAECs were cultured in vitro. The pcDNA3- ICAM-2-Enhancer-CD59cDNA was transfected into the PAECs by lipofectinnmine mediated method. Normal PAECs served as negative control and human PBMCs as positive control. Stable transfectants were selected using G418 （concentration 100 mg/L）. The CD59 protein expression in transfected cells was assayed by flow cytometry. CD59 gene fragments were transferred into mouse zygote by micro-injection and new born pups were detected for transgene and genotyped. Results The expression intensity of pcDNA3- ICAM-2-Enhancer- CD59cDNA recombinant plasmid in PAECs was 63.1%. The expression intensity on the membrane of monocytes in periphery blood of transgenic mice of CD59 gene was 73.38%. Conclusion Recombinant CD 59 protein using ICAM-2 promoter with enhancer can be expressed highly, effectively and specifically in PAECs and transgenic mice.