兔出血症病毒VP60抗原优势区和Th细胞表位融合蛋白的原核表达及免疫效力测定  

Expression of dominant antigen regions of RHDV VP60 capsid fused with universal Th cell epitope and evaluation immune efficacy in rabbits

作　　者：孟春春[1] 程英杰[1,2] 李传峰[1] 王玢瑸[3] 缪秋红[1] 陈宗艳[1] 吴润[2] 刘光清[1] 

机构地区：[1]中国农业科学院上海兽医研究所,上海200241 [2]甘肃农业大学动医医学院,甘肃兰州730070 [3]西北农林科技大学动物医学院,陕西杨凌712100

出　　处：《中国预防兽医学报》2014年第3期239-242,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：农业公益性行业科研专项课题(201303046;201003012);"863"计划(2011AA10A209);国家自然科学基金(C010802)

摘　　要：为检测兔出血症病毒(RHDV)衣壳蛋白VP60两个抗原优势区串联通用Th细胞表位的融合蛋白AB-Th的免疫效力,本研究采用融合PCR的方法以pBL-RHDV为模板扩增AB-Th基因并亚克隆至原核表达载体pET-30a(+)中,转化大肠杆菌BL21(DE3)菌株,大量表达AB-Th融合蛋白。结果显示,融合蛋白以包涵体形式表达,并且具有良好的免疫原性。使用纯化后的融合蛋白免疫实验兔,并分别利用ELISA方法和MTS法检测兔免疫后的抗体水平和淋巴细胞增殖能力。结果表明该融合蛋白能够使兔产生针对RHDV VP60蛋白的特异性抗体和导致高水平的淋巴细胞增殖,并能够为实验兔提供100%的保护。To evaluate the immune efficacy of the dominant antigen regions of Rabbit hemorrhagic disease virus VP60 capsid fused with universal Th cell epitope. The AB-Th gene was cloned into expression vector pET-30a by overlapping PCR. After transformed into E.coil BL21, 1 mmol/mL of IPTG was added to induce the recombinant protein expression. The target protein was purified by gradients of urea and vaccinated rabbits. In all cases, anti-RHDV antibodies were detected by ELISA. Additionally, the lymphocyte proliferation response was tested by the 3-（4,5-dimethyl-2-thiazolyl）-2,5-diphenyl-2H-tetrazolium bromide method. Our results showed that RHDV-specific antibodies and an RHDV-specific cell-mediated immune response were strongly induced in rabbits. Furthermore, all of the rabbits were protected against challenge with wild type RHDV. In conclusion, we demonstrated that the suicidal DNA vaccine is a promising vaccine candidate that facilitates the prevention of rabbit hem-orrhagic disease caused by RHDV.

关 键 词：VP60优势抗原区 Th细胞表位 原核表达 免疫效力测定 

分 类 号：S852.65[农业科学—基础兽医学]