造血器官坏死病病毒M蛋白的可溶性表达及免疫原性分析  

Soluble Expression and Immunogenicity Analysis of M Protein of Hematopoietic Necrosis Virus

作　　者：郑园园 杨延超 李清扬 赵宝华[1] Zheng Yuanyuan;Yang Yanchao;Li Qingyang;Zhao Baohua(College of Sciences,Hebei Normal University,Shijiazhuang,Hebei 050700,China)

机构地区：[1]河北师范大学生命科学学院,河北石家庄050700

出　　处：《中国动物检疫》2023年第3期118-124,共7页China Animal Health Inspection

基　　金：河北省冷水鱼绿色高效养殖技术集成与示范项目(21326703D)。

摘　　要：为在大肠杆菌中高效表达出可溶性的传染性造血器官坏死病病毒(IHNV)基质蛋白M,并分析其免疫原性,构建了pMAL-c2x-m原核表达质粒,优化表达条件并纯化MBP-M融合蛋白;对蛋白进行Western blot和DDA鉴定;将鉴定后的蛋白免疫BALB/c小鼠,利用Western blot鉴定鼠抗MBP-M血清,通过间接ELISA检测该血清效价。结果显示:在大肠杆菌中获得了可溶性的MBP-M蛋白,其相对分子质量约为64 kDa,与预期结果一致;MBP-M蛋白在IPTG为0.5 mmol/L,25℃诱导12 h时上清表达量较高;经Western blot鉴定,MBP-M蛋白可与鼠抗血清发生特异性反应,而不与阴性对照血清发生反应;ELISA检测血清效价达1:25600。结果表明,本研究获得的MBP-M蛋白具有较好的免疫原性,这为IHNV基因工程疫苗研发奠定了基础。In order to express soluble matrix protein M of infectious hematopoietic necrosis virus(IHNV-M)from Escherichia coli(E.coli)and to analyze its immunogenicity,a prokaryotic expression plasmid was constructed and named as pMAL-c2x-m,the expression conditions was optimized and then MBP-M fusion protein was purified.The purified protein was identified by Western blot and DDA,and then was used to immunize BALB/c mice,and the mouse antiserum was identified by Western blot,whose titer was determined by indirect ELISA.The results showed that soluble MBP-M protein was obtained from E.coli,and its relative molecular mass was about 64 kDa as expected;the expression level of MBP-M protein in supernatant was higher when IPTG was 0.5 mmol/L and the bacteria solution was induced at 25℃for 12 h;as identified by Western blot,MBP-M protein could specifically react with mouse antiserum,but failed with negative control serum;and the titer of serum antibodies reached 1:25600 by indirect ELISA.In conclusion,the immunogenicity of the obtained MBP-M protein was good,which laid a foundation for future researches and development of genetic engineering vaccines of IHNV.

关 键 词：传染性造血器官坏死病病毒 麦芽糖结合蛋白 可溶性表达 免疫原性分析 

分 类 号：S852.65[农业科学—基础兽医学]