牛病毒性腹泻病毒E2蛋白的亚细胞定位研究

Subcellular Localization Analysis of Bovine Viral Diarrhea Virus E2 Protein

关键词:  牛病毒性腹泻病毒  E2蛋白  亚细胞定位

Key words:  bovine viral diarrhoea virus  E₂ protein  subcelluar localization

摘要:

为了了解E2蛋白在哺乳动物细胞中的亚细胞定位情况。采用RT-PCR扩增牛病毒性腹泻病毒Cangchun184株E2基因,将其克隆至真核表达载体pcDNA3.1/V5HisA中,并进行酶切鉴定和测序分析,将构建的重组质粒pcDNA3.1/V5HisA-E2经脂质体介导转染MDBK细胞48 h后,在激光共聚焦显微镜下观察E2蛋白在细胞中的亚细胞定位情况。结果表明,E2蛋白在细胞核上和细胞质中均有表达,分布均匀,且绿色荧光大多呈点状均匀分布,为进一步研究牛病毒性腹泻病毒奠定基础。

Abstract:

For further research on subcellular localization of BVDV E₂ protein in mammalian cells. E₂ gene fragments of Changchun184 were amplified by RT-PCR and cloned to eukaryotic expression vector pcDNA3.1/V5HisA. Through identification of enzyme digestion and sequencing, the recombinant plasmid pcDNA3.1/V5HisA-E₂ was constructed and transfected into MDBK cells by liposome mediated .After transfection 48h,subcellular location of E₂ protein in MDBK cells was observed by confocal laser scanning microscope. The results showed that E₂ protein was expressed and distributed in both the nucleus and cytoplasm, and green fluorescence was mostly punctate distribution. These results laid the foundation for further study to bovine viral diarrhea virus.