不同致病型的三株IBV与chAPN体外结合能力的比较  

Comparison of the Binding Capacity on three Pathotype IBV Strains with Chicken Aminopeptidaes N in vitro

作　　者：刘澜澜[1] 明晓波[2] 徐放[1] 王亚贤[1] 魏萍 

机构地区：[1]黑龙江中医药大学基础医学院,黑龙江哈尔滨150040 [2]东北农业大学动物医学院,黑龙江哈尔滨150030

出　　处：《现代生物医学进展》2015年第17期3243-3246,共4页Progress in Modern Biomedicine

基　　金：黑龙江中医药大学校科研基金(BS201402);黑龙江省教育厅科学技术研究项目(2003fz012)

摘　　要：目的:比较不同致病型鸡传染性支气管炎病毒(Infectious bronchitis virus,IBV)IBV与鸡氨肽酶N体外结合能力的差异。方法:将已构建的p ET-32a-ch APN转入大肠杆菌BL21中进行诱导表达,SDS-PAGE分析表达结果。表达的His-ch APN纯化后进行western-blotting验证,应用酶促反应检测其活性。以不同稀释度的纯化ch APN包被ELISA板,加入经过荧光定量PCR方法定量的含有相同数量病毒粒子的三株不同致病型IBV(呼吸型M41、肾型S-03和腺胃型A2-2),测定三株与ch APN的结合能力。结果:p ET-32a-ch APN在受体菌内成功表达,主要以包涵体的形式存在。经纯化复性的重组His-ch APN,Western blotting分析显示其具有良好的抗原性,酶促反应显示该重组重组His-ch APN与天然的具有良好的生物活性。ELISA结果显示ch APN与三株IBV均能结合,其结合能力为A2-2>S-03>M41,并表现为剂量依赖性。结论:不同致病型IBV与ch APN的体外结合力有一定差异。Objective： To compare the binding capacity on different pathotype infectious bronchitis virus（IBV） strains with cellular protein chicken Aminopeptidase N（ch APN） in vitro. Methods： The p ET-32a-ch APN was converted into E.coli BL21 cells to induce expression. SDS-PAGE analysis were used to examine the fusion protein. The purified His-ch APN was detected by Western blotting and its activity was determined by enzymatic reaction. ELISA plates were coated by the different dilution His-ch APN, and the equivalent amount of three different pathotype IBV strains（respiratory type IBV strain M41, nephropathogenic IBV strain S-03 and proventriculitis type IBV strain A2-2） quantified by Q-PCR were added into the wells, determined the difference among the binding capacity on three different pathotype IBV strains and ch APN in vitro. Results： It showed that the recombinant p ET-32a-ch APN was expressed successfully in an inclusion body form. The purified His-ch APN had expected antigenicity tested by western blotting analysis and had enzymatic activity which was similar to the natural ch APN. ELISA data demonstrated the His-ch APN was able to bind three IBV strains dose-dependently, A2-2S-03M41. Conclusions： The binding capacity on different pathotype IBV strains with ch APN in vitro was different to a certain extent.

关 键 词：致病型 IBV CH APN 结合能力 

分 类 号：Q95-3[生物学—动物学] S858.31[农业科学—临床兽医学]