含NDV F48E9株F基因的MDV CVI988株转移质粒载体的真核表达  

Construction and eukaryote expression of marek's disease virus CVI988 transferring vector containing NDV F gene

作　　者：苏春霞[1] 苏鑫铭[1] 张彦明[2] 曹瑞兵[1] 周斌[1] 陈溥言[1] 

机构地区：[1]南京农业大学农业部畜禽疫病诊断与免疫重点开放实验室,江苏南京210095 [2]西北农林科技大学动物科技学院,陕西杨凌712100

出　　处：《西北农林科技大学学报（自然科学版）》2006年第5期59-62,共4页Journal of Northwest A&F University(Natural Science Edition)

摘　　要：为进一步提高重组病毒的表达水平,用构建的含gB启动子和NDV F48E9株F基因的MDV CVI988株转移质粒载体pUS10F转染293T细胞,通过PCR和接免疫荧光(IFA)法研究了其在真核中的表达,并确定最佳的转染条件。结果表明,真核细胞中带有F基因,转移质粒载体已经进入细胞DNA;转染转移质粒载体 pUS10F后的293 T细胞所表达的NDV F基因产物抗原性较好,能够与抗NDV的标准血清反应。确定的最佳转染条件为:DNA 3μg／mL,转染后48 h外源基因在细胞中的表达量最高,在6孔板和96孔板中细胞数量分别以 2×105／孔和1×101／孔的转染效果最好。In order to improve expression level of recombinant virus,the transferring vector pUS10F with gB promotor of MDV and F gene of NDV was transfected in 293T cells. Expression of the gene expression cassette in the cells was detected by PCR and IFA,and the transferring condition was optimized. The result shows that gene cassette was integrated in the DNA of cells. Expression products present good antigenicity because it can react with anti-NDV standard serum. The best transferring condition is ：DNA 3 μg/mL;the highest expression level was testified 48 h after gene was transfected 293T cell;the best transfected effect was proved in 6-wells micro titer plate,at 2 × 10^5 cells/well and in 96-wells micro titer plate,at 1 × 10^4 cells/well.

关 键 词：畜禽疫病 新城疫病毒 F基因 马立克氏病病毒CVI988转移质粒载体 

分 类 号：S858.312.659.5[农业科学—临床兽医学] S858.312.659.1[农业科学—兽医学]