猪IL-15 cDNA真核表达载体的构建、表达及其免疫佐剂效应  

Construction and expression of porcine interleukin-15 cDNA eukaryotic expression vector and its enhancing of immunity as immunoadjuvants

作　　者：何谦[1] 陈钜豪[2,3] 房红莹[3] 卢俊鹏[3] 张欣[3] 崔敏[3] 曾小娜[3] 刘健[3] 罗满林[3] 

机构地区：[1]暨南大学生命与健康工程研究院,广东广州510632 [2]广州市工商行政管理局白云分局,广东广州510402 [3]华南农业大学兽医学院动物传染病实验室,广东广州510642

出　　处：《中国兽医学报》2011年第8期1147-1151,共5页Chinese Journal of Veterinary Science

摘　　要：参考GenBank发表猪IL-15mRNA序列设计引物,用RT-PCR方法扩增猪IL-15cDNA,并克隆到pMD18-T载体中,通过PCR、酶切和测序验证克隆正确,再亚克隆到真核表达载体pcDNA-3.1(+)上,得到重组质粒pcD-NA-pIL-15。在脂质体介导下,重组质粒pcDNA-pIL-15转染AD-293细胞。以鼠抗猪IL-15为一抗,用间接免疫荧光分析表明猪IL-15基因均在AD-293细胞中成功进行了瞬时表达。小鼠免疫试验表明,pcDNA-pIL-15作为免疫佐剂,能够有效提高小鼠的脾T细胞增殖,加强pcDNA-ORF2(PCV2)质粒免疫过程中的特异性中和抗体的产生,为进一步研制猪IL-15基因佐剂疫苗及进行临床实验奠定基础。According to the sequences of porcine interleukin-15 messenger ribonucleic acid（pIL-15mRNA） published in GenBank,the primers were designed to amplify IL-15 cDNA,which was cloned to into the pMD18-T vector.The product was proved correctly by PCR,enzyme digestion and sequencing,and subcloned into the eukaryotic expression vector pcDNA-3.1（＋）,then the recombinant plasmid pcDNA-pIL-15 was transfected to AD-293 cells by the liposome-mediated.With mouse anti-pig IL-15 as the first antibody,indirect immunofluorescence analysis showed that the porcine IL-15 gene were successfully carried out and transiently expressed in AD-293 cells,The immunization test to Balb/c mice showed that as an immunologic adjuvants,the pcDNA-pIL-15 could promote the proliferation of T lymphocyte cells and enhanced the specific antibody level for pcDNA-ORF2（PCV2） plasmids,which establish the foundation for further development of pig IL-15 gene adjuvant vaccine and conduct clinical trials.

关 键 词：猪IL-15 真核表达 瞬时表达 佐剂疫苗 中和抗体水平 

分 类 号：S828[农业科学—畜牧学]