转EGFP基因山羊克隆胚的发育

利用脂质体包裹含EGFP基因的质粒,并将之导入山羊乳腺上皮细胞,经G418筛选获得阳性细胞,以阳性细胞作为核供体,利用核移植技术构建转基因克隆胚。结果表明：用转基因山羊乳腺上皮细胞作为核供体,电融合法更适合构建转基因克隆胚。转基因山羊克隆胚体外培养最佳方案是用SOFaa培养液,在培养72 h后加入10%的正常山羊血清（Normal goat serum,NGS）。荧光显微镜下观察到转基因克隆胚中EGFP的表达,大部分克隆胚发育到8-16细胞以后的时期,绿色荧光蛋白才开始逐渐表达,随着发育时间的延长,绿色荧光蛋白的表达也逐渐增强。说明外源基因在胚胎早期发育阶段可以表达,EGFP可以作为报告基因来实现对外源基因整合及表达的监测。

Development of Goat Cloned Embryos Containing EGFP Gene

Plasmid containing EGFP gene was transfected into the goat mammary gland epithelial cells by lipofection.Stable transfectants were selected by G418 and the positive cells were used as nucleus donors to create transgenic goat cloned embryos by nuclear transfer techniques.SOFaa culture medium supplied with 10 % normal goat serum(NGS) was the best combination for culturing the transgenic goat cloned embryos in vitro.Expression of the foreign gene in the embryos was detected by observation of green fluorescence.EGFP was expressed gradually after the 8-16 cell stages in the most of embryos,and the level of EGFP expression increased with development of the embryos.The results implied that EGFP gene could be used as reporter gene to check expression of foreign gene in the transgenic goat cloned embryos in early stages.