鸡传染性贫血病毒vp1基因的克隆表达及抗原性分析  

Expression and antigenic analysis of chicken anemia virus vp1 genes

作　　者：杨勇[1] 任玉红[1] 梁宝怀 朱雅宁[3] 梁志选[3] 

机构地区：[1]山西农业大学动物科技学院,山西太谷030801 [2]山西省大同市南郊区畜牧局,山西大同037001 [3]天津市禽病诊断培训中心,天津300402

出　　处：《中国预防兽医学报》2007年第8期575-578,583,共5页Chinese Journal of Preventive Veterinary Medicine

摘　　要：从组织病料中提取CAV核酸,根据GenBank上发表的序列设计两对引物,分别扩增出vp1的139 bp～83 bp和547 bp～1337 bp两个基因片段,然后分别将其充隆到原核表达载体PMXB10上,经PCR和酶切鉴定,证明成功构建了重组表达载体PMXB10-vp1C和PMXB10-vp1D。在0.3 mM的IPTG诱导下,融合蛋白在大肠杆菌ER_(2566)以分泌型得到大量表达,经大量表达后,用几丁质柱挂柱切割纯化后,得到切割蛋白E、F,在Western blot免疫分析印迹中,两组融合蛋白和切割蛋白与CAV阳性血清均能发生特异性反应。用ELISA方法检验,纯化的两组蛋白与CAV阳性血清均能发生特异性反应。用两组蛋白免疫SPF鸡后,用全病毒ELISA试剂盒检测血清呈阳性,表明两组蛋白均可诱发机体产生抗CAV的抗体。Two CAV vpl gene fragments of 139 bp-830 bp and 547 bp-1 337 bp were amplified from virus DNA by PCR using primers designed according to published sequences. The PCR products were cloned into prokaryotic expression vector PMXB10, producing recombinant expression vector PMXB10-vplC and PMXB10-vpl D. The resultant constructs were transformed into E.coli ER2566 cells and proteins were expressed by IPTG induction. The expressed proteins were determined by Western blot. Both proteins reacted specifically with CAV positive serum in ELISA. Serum of SPF chicken immunized with the recombinant proteins was shown to react positively with whole virus in ELISA, indicating that the proteins induced antibodies against CAV.

关 键 词：鸡传染性贫血病毒 VP1 原核表达 酶联免疫吸附实验 

分 类 号：S852.659.2[农业科学—基础兽医学] Q786[农业科学—兽医学]