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Cos7细胞后的表达产物,可利用抗HA或myc的mAb进行Westernblot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。
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.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2001年第2期175-178,共4页
Chinese Journal of Cellular and Molecular Immunology
基金
上海市现代生物与新药产业发展基金资助No.975419001-1
