水产学报2011，Vol.35Issue(8)：1166-1171,6.DOI:10.3724/SP.J.1231.2011.17315
刺参甘露糖结合凝集素的原核表达、纯化及生物活性分析
Prokaryotic expression, purification and bioactivity analysis of Apostichopus japonicus mannan-binding lectin
摘要
Abstract
To further study the functional characteristics of Apostichopus japonicus mannan-binding lectin ( AJ-MBL)and to improve the natural immunity of A. Japonicus,a prokaryotic vector of AJ-MBL gene CDS region was constructed. The fusion protein was expressed and purified in prokaryotic system and its bioactivity was studied. The coding sequence of AJ-MBL was amplified by RT-PCR method. After being identified by the restriction digestion and sequencing, the 498 bp of AJ-MBL gene was inserted into pET28a plasmid to yield an identified recombinant plasmid Pet-AJ-MBL, which was used to transform the competent expressive cells of E. Coli BL21(DE3). After induction with IPTG,samples analysis results revealed that a fusion protein of approximately 17 ku was yielded,and it occurred in the form of inclusion bodies,and could be purified with Ni2+ affinity chromatography. The results showed highly expressed fusion protein was acquired. The purified 17-ku AJ-MBL underwent hemagglutination assay to test its bioactivity, the results showed that the minimum hemagglutination concentration was 10 (xg/Ml. These results indicated that the CDS domain of AJ-MBL had high expressed and the fusion protein(17 ku AJ-MBL) was highly bioactivity.关键词
刺参/甘露糖结合凝集素/原核表达/蛋白纯化/红细胞凝集试验Key words
Apostichopus japonicus/ mannan-binding lectin/ prokaryotic expression/ purification/ hemagglutination assay分类
生物科学引用本文复制引用
李丹彤,谢广成,丁文勇,王秀利,刘洋,徐文琦,张永攀..刺参甘露糖结合凝集素的原核表达、纯化及生物活性分析[J].水产学报,2011,35(8):1166-1171,6.基金项目
辽宁省教育厅团队项目(2008T021) （2008T021）
