禽白血病病毒gp85蛋白在293T细胞中的表达及分析

Expression and Analysis of Avian Leukosis Virus gp85 Protein in 293T Cells

为获得293T细胞表达的ALV-J gp85蛋白,研究通过PCR扩增ALV-J gp85基因,利用基因克隆技术克隆pMD-18T-gp85并测序,进而构建表达质粒p EGFP-N1-gp85。利用293T细胞对重组表达质粒进行表达,通过荧光显微镜和Western blot检测重组表达质粒(pEGFP-N1-gp85)的表达情况。结果显示,pEGFP-N1-gp85重组表达质粒在293T细胞中均匀分布,说明pEGFP-N1-gp85重组表达质粒在293T细胞中表达;pEGFP-N1-gp85重组表达质粒在293T细胞中表达的蛋白质分子质量约为60 ku。结果表明,成功构建了真核表达质粒pEGFP-N1-gp85,并在293T细胞中获得了表达,为研制ALV-J-gp85 DNA核酸疫苗提供了科学支撑。

In order to obtain the ALV-J gp85 protein expressed in 293 T cells, the ALV-J gp85 gene was amplified by PCR, and pMD-18 T-gp85 was cloned and sequenced by gene cloning technique, and the expression plasmid pEGFP-N1-gp85 was cloned. The recombinant expression plasmid was expressed by human renal epithelial cell line293 T cells, and the expression of recombinant expression plasmid was detected by fluorescence microscopy and Western blot. The results showed that pEGFP-N1-gp85 recombinant expression plasmid was evenly distributed in 293 T cells,indicating that pEGFP-N1-gp85 recombinant expression plasmid was expressed in 293 T cells.The recombinant expression plasmid expressed a positive band at a protein molecular mass of about 60 ku in 293 T cells. The results indicated that the eukaryotic expression plasmid pEGFP-N1-gp85 was successfully constructed and expressed in 293 T cells,which provided scientific support for the development of ALV-J-gp85 DNA nucleic acid vaccine.