H5N1亚型禽流感病毒NP蛋白一个鸡MHCⅠ分子限制性T细胞表位的鉴定  

Identification of the MHC class I-restricted cytotoxic T cell epitope in the H5N1 avian influenza virus nucleoprotein

作　　者：朱凤珠 许传田[2] 鲁梅[3] 张秀美[2] 黄庆华[2] 黄艳艳[2] 杨少华[2] 吴家强[2] 谭刘刚 崔言顺[1] 

机构地区：[1]山东农业大学动物科技学院,山东泰安271018 [2]山东省农业科学院畜牧兽医研究所山东省畜禽疫病防治与繁育重点实验室,山东济南250100 [3]山东潍坊工程职业学院,山东青州262500

出　　处：《中国预防兽医学报》2015年第11期861-865,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：现代农业产业技术体系(CARS-42-Z12);公益性行业(农业)科研专项(201303033);十二五国家科技支撑计划(2015BAD12B03)

摘　　要：为鉴定禽流感病毒核蛋白(NP)中的T细胞表位,本研究根据蛋白结构预测技术预测了4个潜在细胞毒性T淋巴细胞(CTL)表位(NP120-128、NP163-172、NP85-92、NP67-74),并合成相应的候选短肽。构建了含有NP基因的重组质粒p CAGGS-NP,将其转染293T细胞。间接免疫荧光和western blot结果表明NP蛋白能够在293T细胞中获得表达。重组质粒免疫SPF鸡后能够诱导鸡体产生特异的NP抗体。在合成的4条短肽中,NP67-74体外刺激免疫过p CAGGS-NP的鸡脾淋巴细胞可以显著提高脾淋巴细胞IFN-γ的分泌量,提示短肽NP67-74能够在体外诱导活化鸡淋巴细胞产生CTL,可能是AIV NP的CTL表位。该研究结果对流感病毒免疫机制和通用疫苗研究具有借鉴意义。In order to identify the T cell epitope in avian influenza virus （A1V） nucleoprotein （NP）, four T cell epitope candidate peptides （NP104-112 , NP163-172, NP85-92 and NP67-74） of the AIV NP were predicted with the protein structure prediction technology and synthesized. To verify the immunogenicity of these peptides, the recombinant plasmid pCAGGS-NP was constructed and the expression of NP was confirmed by indirect immunofluorescence and western blot in transfected 293T cell. The antibodies against NP were also detected by ELISA in pCAGGS-NP inoculated SPF chickens. The splenic cells were collected from inoculated chickens and stimulated with the peptides of NP120-128, NP163-172, NP85-92, NP67-74 or negative control, respectively. The results of real-time PCR show that spleen lymphocytes stimulated with the peptide NP67.74 significantly increased the IFN-7 secretion, which indicated that the peptide NP67-74 might be AIV NP CTL epitope. The results possessed significance to mechanism of protective immunity against influenza virus and the research towards universal flu vaccine.

关 键 词：禽流感病毒 NP蛋白 CTL表位 

分 类 号：S852.65[农业科学—基础兽医学]