牛泡沫病毒BFV3026细胞感染性的确立及包装细胞系的建立

分别将牛泡沫病毒BFV3026接种于胎牛肺细咆(FBL）、牛肺细咆系(BL12)、新生牛肾细胞(NBK)、兔肺细胞(RL)、人乳腺癌上皮细咆(MCF)、293T、HeLa、CV-1、CHO等9种细胞，通过对它们及其传代细胞的病变观察-RT—PCR检测，确立BFV3026对这9种细胞的感染。并以BFV3026原病毒DNA为模板，通过PCR构建以pcDNA3．1(-)为载体的gag—pol、env真核表达质粒共转染BL12细胞，经G418持续筛选，获得8个Neo^R细胞克隆RT—PCR及包装实验证实：其中7个细胞克隆能有效行使包装辅助功能。

Characterization of BFV3026 Infection and Construction of the Packaging Cell Line

Fetal bovine lung cell(FB L),bovine lung cell(BL12),new born bovine kidney cell (NBK),rabbit lu ng cell(RL),CV-1,MCF,293T,HeLa and CHO cells were infected with bovine foam y virus(BFV3026).Virus infection i n these 9 cell lines was confirmed by detection of the cytopathic eff ects(syncytium formation)and BFV internal promoter(IP)regi on by RT-PCR in the infected cells and their passaged cells.In addition,gag-pol and env genes were cloned from the full-length DNA of BFV3026 and use d to co-transfect BL12.Stable bovi ne packaging cell line had been ge nerated by subjecting transfectant s to neomycin selection and screen ing for coincident expression of t hree structure genes using RT-PCR. The capacity of package was measur ed with transfecting of BFV transf er vehicle into the packaging cell lines and infecting of the recover ed defective viruses into fresh B L12 cells,which contain luciferase report gene cloned under the contr ol of BFV IP.