论文部分内容阅读
目的 构建人嗜酸性粒细胞来源的神经毒素(hEDN)与乙肝病毒核心蛋白 (HBVc)的融合真核表达载体(该融合蛋白即为构建的靶向核糖核酸酶 ) ,抑制乙肝病毒在细胞内的复制。方法 分别从HL 6 0细胞和 2 2 15细胞中提取总RNA ,用RT PCR特异性扩增hEDN及HBVc编码基因 ,将hEDN和HBVc克隆入真核表达载体pcDNA3 1(- ) ,hEDN克隆入原核表达载体 pGEX4T 1,并以纯化的原核表达产物免疫Balb/c小鼠 ,制备特异性抗体。用免疫荧光法检验 pcDNA3 1(- ) /HBVc -hEDN在转染 2 2 15细胞内地表达。结果 成功地构建了hEDN和HBVc的真核融合表达载体 ,并在 2 2 15细胞中较高效地表达。结论 pcDNA3 1(- ) /HBVc hEDN的构建和在 2 2 15细胞内的表达 ,为探索乙型肝炎的治疗开辟了新思路
Objective To construct a fusion eukaryotic expression vector of human eosinophil derived neurotoxin (hEDN) and hepatitis B virus core protein (HBVc), which is a targeted ribonuclease, and inhibit the intracellular copy. Methods The total RNA was extracted from HL 60 cells and 2 215 cells respectively. The hEDN and HBVc genes were amplified by RT PCR. The hEDN and HBVc were cloned into the eukaryotic expression vector pcDNA3 1 (-) and the hEDN was cloned into prokaryotic The expression vector pGEX4T 1 was used to immunize Balb / c mice with the purified prokaryotic expression product to prepare specific antibodies. The expression of pcDNA3 1 (-) / HBVc-hEDN was detected by immunofluorescence in transfected 2 215 cells. Results The eukaryotic fusion expression vector of hEDN and HBVc was successfully constructed and expressed more efficiently in 2 15 cells. Conclusion The construction of pcDNA3 1 (-) / HBVc hEDN and its expression in 2 215 cells opened up new ideas for exploring the treatment of hepatitis B