猪繁殖与呼吸综合征病毒HB-3株GP5-M-N蛋白核酸疫苗的构建及免疫原性  

Construction and immunogenicity of recombinant plasmid co-expressing GP5,M and N proteins of porcine reproductive and respiratory syndrome virus

作　　者：杜冬华 周静[1] 王爱华[1] 陈赛娟[2] 刘涛 

机构地区：[1]河北北方学院动物科技学院,河北张家口075131 [2]河北农业大学,河北保定071000 [3]保定瑞普生物药业有限公司,河北保定071000

出　　处：《中国兽医学报》2014年第2期177-181,共5页Chinese Journal of Veterinary Science

基　　金：河北省重大技术创新基金资助项目(07221008Z)

摘　　要：为探索猪繁殖与呼吸综合征病毒(PRRSV)核酸疫苗用于免疫预防的可行性,试验用PCR方法扩增出PRRSV HB-3株GP5、M和N基因,通过Linker序列将GP5和M串联为GP5-M,双酶切后和N基因一起插入真核表达载体构建重组质粒pcDNA-GP5-M-N,经酶切鉴定表明GP5、M和N基因和载体连接正确。然后将重组质粒转染至Marc-145细胞,经间接免疫荧光及Western blot分析证实重组蛋白能在Marc-145细胞中表达。然后用重组质粒pcDNA-GP5-M-N免疫Balb/c小鼠,中和抗体检测结果表明,首免后2周即有小鼠产生可检测到的病毒中和抗体(1∶4),随后抗体水平快速升高,第8周抗体效价达到最高(1∶32)。说明本试验构建的重组质粒pcDNA-GP5-M-N能诱发免疫小鼠产生较高水平的中和抗体,为PRRSV核酸疫苗的研究奠定了基础。This expieriment was conducted to study the DNA vaccine for preventing PRRSV infection. The GP5,M and N genes of porcine reproductive and respiratory syndrome virus （PRRSV） were amplified by PCR, and then were subcloned into eukaryotic expression vectors pcDNA3. 1 （＋）. The recombinant named as pcDNA-GP5-M-N was transfected into Marc-145 cells,and its ex pression was detected by indirect immune fluorescence detection. The results showed that GP5 M- N genes of PRRSV HB-3 strain were expressed in Marc-145 cells. Using pcDNA-GP5-M-N to im munize Balb/c mice,anti-PRRSV neutralizing antibodies were detected at two weeks after primary vaccination and reached a peak of titer （1 = 32） at eight weeks. These results showed that recombi- nant plasmid pcDNA-GP5-M-N has been constructed successfully,and with good immunity. These data indicated that pcDNA-GP5-M-N could be used for further development of DNA vaccines for PRRSV.

关 键 词：猪繁殖与呼吸综合征病毒 GP5 M N基因 核酸疫苗 免疫原性 

分 类 号：S852.65[农业科学—基础兽医学]