【摘要】 目的构建含有HA和myc编码序列的真核细胞融合表达载体,用于验证两种蛋白质之间的相互作用。方法利用寡核苷酸合成和PCR技术,扩增HA和myc的部分编码序列。在哺乳动物细胞表达载体pcDNA3的基础上,构建了pcDNA3HA和pcDNA3myc两种真核融合表达载体。结果将两种候选蛋白的编码序列分别克隆于上述融合载体,转染Cos7细胞后的表达产物,可利用抗HA或myc的mAb进行Westernblot和免疫共沉淀。结论利用上述系统,可验证两种候选蛋白在哺乳动物细胞中是否具有相互作用。更多还原

【Abstract】 Aim To construct two fusion vectors carrying gene coding HA or myc. Methods Based on the mammalian expression vector pcDNA3, two fusion vectors pcDNA3(HA) and pcDNA3(myc) were constructed by oligonucleotide synthesis and PCR amplification. Results The sequences coding two target proteins were cloned into the above recombinant expression plasmids separately and transfected into Cos 7 cells mediated via liposome. The expression products were detected by Western blot and co immunoprecipitation. Conclusion A system for validating the interactions between two proteins in mammalian cells is established.更多还原