【摘要】 为了探讨影响山羊体细胞中外源基因整合效率的因素,采集胎羊和小羊皮肤并制备成纤维细胞,采用"Lipofectamine LTX"脂质体转染试剂包埋外源基因,分别导入上述细胞中,比较同一种外源基因载体(人凝血因子Ⅸ)转染至不同个体和类型的羊细胞,以及不同载体(人凝血因子Ⅸ、人转铁蛋白和牛催乳素)转染至同一个体羊细胞的整合效率的差异.通过药物筛选得到单细胞簇,经定量PCR计算外源基因的整合效率.同一种外源目的基因载体转染小羊成纤维细胞的整合效率显著高于胎羊成纤维细胞(P<0.05);而不同目的基因载体转染同一个体羊细胞,其细胞的整合效率与转染载体的片段大小有关.由此可知,小羊成纤维细胞外源基因整合频率优于胎羊细胞,且载体片段的大小影响外源基因转染的整合频率.更多还原

【Abstract】 To investigate the impacts of foreign DNAs in goat somatic cells on the integration efficiencies,the skins were collected from fetal and young goats and the fibroblasts were then cultured.The foreign gene vectors were transfected into the goat fibroblasts using"Lipofectamine LTX",and the integrated frequencies were com-pared between the cells derived from different types of goats transfected with the same foreign vector(hFIX) as well as between the cells derived from the same goat but transfected with three different vectors(hFIX,hTF or bPRL).The monocloned cell clusters were obtained with the drug selection and the integrated frequency was calculated with qPCR.The integration efficiency of the young goat fibroblasts was higher than the fetal cells(P<0.05).In the same cells transfected with different foreign genes,the integration frequencies were relevant with the length of the vectors.In conclusion,the integrated efficiency in young goat fibroblasts was better than in the fetal cells for transfected the same foreign gene.In addition,the sizes of the DNA fragments transfected had the effect on integration efficiency.更多还原