摘要
IL-18作为一种多效性细胞因子,在机体免疫应答及各种生理功能中发挥着重要的调节作用,根据猪IL-18基因序列设计1对引物,将编码猪IL-18的成熟蛋白基因亚克隆到杆状病毒转移载体pFastBacDual中,并在C端融合6个组氨酸标签以利于纯化,然后转化到含穿梭载体Bacmid的感受态细胞DH10Bac中,发生转座作用。将重组质粒转染昆虫细胞,SDS-PAGE可检测到分子量为18 kDa左右的重组蛋白,Western blotting证实该重组蛋白可与兔抗猪IL-18抗体发生特异性反应。纯化后蛋白能明显促进猪T淋巴细胞转化,表明所表达的IL-18具有较高的生物活性。此研究为进一步开发研制新型免疫佐剂奠定了基础。
IL-18,as a polyphonic cytokine,is important in immune response and physiologic function.We designed one pair of primers,amplified the porcine IL-18 gene fused with a C-terminal 6×Histidine tag,and then subcloned into the pFastBacDual of Baculovirus transfer vector and transformed into DH10Bac containing a shuttle vector of Bacmid.After co-transfecting the recombinant plasmid into insect cells,the 18 kDa expressed protein of porcine IL-18 was detected by SDS-PAGE;the specificity of expressed protein was confirmed by Western blotting.The purified porcine IL-18 protein induced obvious proliferation of porcine T lymphocytes in vitro,which indicated that the expression of IL-18 had high biological activity.
出处
《生物工程学报》
CAS
CSCD
北大核心
2011年第1期118-123,共6页
Chinese Journal of Biotechnology
基金
国家"十一五"科技支撑计划(No.2006BAD06A08)资助~~
