期刊论文

中文

中国兽医科学

1673-4696

2024

54

05

632-640

10.16656/j.issn.1673-4696.2024.0074

2022YFD1800600:国家重点研发计划

本校为第一机构

本试验旨在建立犬α干扰素(CaIFN-α)原核表达及纯化方法,并对其抗病毒活性进行研究.通过构建pET-28a/CaIFN-α表达载体,高效表达和纯化rCaIFN-α;采用CCK-8 法在犬肾细胞(MDCK)上测定rCaIFN-α 的细胞毒性,在 MDCK/VSV 系统中测定其抗病毒活性效价;RT-qPCR 检测 rCaIFN-α 作用MDCK细胞后下游ISGs的表达水平;进一步利用TCID50 和间接免疫荧光(IFA)测定rCaIFN-α抗犬副流感病毒(CPIV)和犬瘟热病毒(CDV)的感染能力.电泳和Western-blot结果显示,成功制备原核表达的特异性rCaIFN-α;CCK-8 结果显示重组蛋白无明显细胞毒性,细胞病变抑制法测定抗VSV 病毒活性效价为1.39×107 U/mL;RT-qPCR 显示rCaIF...

The aim of this experiment was to establish a method for the prokaryotic expression and purification of canine interferon-α(CaIFN-α)and to research its antiviral activity.rCaIFN-α was ef-ficiently expressed and purified by constructing pET-28a/CaIFN-α expression vector.The cytotoxicity of rCaIFN-α was determined on canine kidney cells(MDCK)by CCK-8 assay,and its antiviral activity po-tency was determined in the MDCK/VSV system.RT-qPCR was used to detect the expression level of down-stream ISGs after rCaIFN-α acted on the MDCK cells.TCID50...