鸡IL-18在毕赤酵母中的表达与活性分析  

Expression and analysis of recombinant chicken IL-18 in Pichia pastoris

作　　者：赵敏[1,2] 张春杰[1] 李银聚[1] 吴庭才[1] 程相朝[1] 刘一尘[1] 俞晓丽[1] 钱永华[2] 

机构地区：[1]河南科技大学动物科技学院,河南洛阳471003 [2]西北农林科技大学动物医学院,陕西杨凌712100

出　　处：《细胞与分子免疫学杂志》2009年第12期1088-1091,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：河南省自然基金项目资助(0511032300)

摘　　要：目的:在毕赤酵母中表达鸡白介素18,并鉴定其活性。方法:应用PCR技术从重组质粒pMD18-T-ChIL-18中扩增出鸡IL-18成熟肽基因,亚克隆于毕赤酵母表达载体pPICZαA上,构建重组质粒pPICZαA-ChIL-18。经酶切、PCR和测序鉴定正确后,电转化入毕赤酵母菌X-33,筛选多拷贝单克隆进行诱导表达。表达产物纯化后用Western blot、ELISA和淋巴细胞转化试验分析其生物学活性。结果:重组菌正确表达了具有免疫原性的鸡IL-18,经分析该rchIL-18具有诱导淋巴细胞分泌γ干扰素和刺激淋巴细胞增殖的活性。结论:成功的在毕赤酵母中表达了具有生物活性的鸡IL-18。AIM： Expression and analysis of recombinant chicken IL-18 in Pichia pastoris. METHODS： Chicken IL-18 mature peptide gene was amplified from the recombinant plasmid pMD18-T-ChlL-18 by PCR, and was subcloned into Pichia pastoris expression vector pPICZαA to construct the recombinant plasmid pPICZaA-ChlL-18. After identified by restriction enzymes digestion analysis, PCR and DNA sequencing, the recombinant plasmid was transformed into Pichia pastoris X-33. Then choosing the multi-copy recombinant strains to be induced for expression. Then the bioactivity of rchIL-18 was analysed by Western blot, ELISA and MTT- after purified by Sephadex G-100 column. RESULTS： The chicken IL-18 with the immunogenicity was secreted by Pichia pastoris. It could induce T lymphocytes proliferation and secreting IFN-y in vitro. CONCLUSION： The chicken IL-18 with obvious biological activity is secreted by Pichia pastoris X-33.

关 键 词：鸡IL-18 毕赤酵母 诱导表达 活性检测 

分 类 号：R392.12[医药卫生—免疫学]