[1]郭松林,陆盼盼,冯建军,等.创伤弧菌OmpU与嗜水气单胞菌OmpⅡ外膜蛋白二联表达及其初步免疫原性[J].集美大学学报(自然版),2017,22(3):1-7.
 GUO Songlin,LU Panpan,FENG Jianjun,et al.Combined Expression and Primary Immunogenicity Study of Outer Membrane Protein OmpU and OmpⅡ of Vibrio vnlnificus and Aeromonas hydrophila[J].Journal of Jimei University,2017,22(3):1-7.
点击复制

创伤弧菌OmpU与嗜水气单胞菌OmpⅡ外膜蛋白二联表达及其初步免疫原性()
分享到:

《集美大学学报(自然版)》[ISSN:1007-7405/CN:35-1186/N]

卷:
第22卷
期数:
2017年第3期
页码:
1-7
栏目:
水产、食品与生物技术
出版日期:
2017-05-28

文章信息/Info

Title:
Combined Expression and Primary Immunogenicity Study of Outer Membrane Protein OmpU and OmpⅡ of Vibrio vnlnificus and Aeromonas hydrophila
作者:
郭松林12陆盼盼12冯建军12林鹏1
(1.集美大学水产学院,福建 厦门 361021;2.鳗鲡现代产业技术教育部工程研究中心,福建 厦门 361021)
Author(s):
GUO Songlin12LU Panpan12FENG Jianjun12LIN Peng1
(1.Fisheries College,Jimei University,Xiamen 361021,China;2.Engineering Research Center of the Morden Industry Technology for Eel,Ministry of Education,Jimei University,Xiamen 361021,China)
关键词:
创伤弧菌迟缓爱德华氏菌OmpUOmpⅡ表达产物免疫原性
Keywords:
Vibrio vulnificusAeromonas hydrophilaompUompⅡexpressionimmunogenicity
文献标志码:
A
摘要:
从发病的养殖鳗鲡中分离出创伤弧菌(Vibrio vulnificus)和嗜水气单胞菌(Aeromona shydrophila),提取其基因组DNA,再通过PCR法克隆创伤弧菌外膜蛋白OmpU和嗜水气单胞菌外膜蛋白OmpⅡ的基因全长。采用融合PCR法体外连接这2个基因表达膜外片段的序列并成功构建了二联表达载体(pGEX-2T-Vibr-Aero-his)。在大肠杆菌(BL21)的吸光度A600为0.6~1.0时,采用1.0 mmol/L IPTG诱导剂,16 ℃过夜诱导表达。表达产物经离心柱亲和层析纯化后获得分子量为82.2 ku的外膜蛋白。蛋白经透析复性后免疫于鳗鲡以测定其血清抗体效价。结果表明,重组蛋白注射组的鳗鲡血清抗体效价在免疫后第14天和第21天显著(P<0.05)高于PBS对照组,第28天和第42天达到极显著(P<0.01)水平。
Abstract:
In this study,template DNAs were extracted from pathogenic Vibrio vulnificus and Aeromonas hydrophila isolated from diseased eels and confirmed by challenge experiment.According to gene sequences coding the protein Omp in GenBank database,two pairs of primers were designed respectively and the whole gene fragments of OmpⅡ of A.hydrophila and OmpU of V.vulnificus were amplified by PCR.Two gene fragments,encoding the out part of the membrane region of OmpU and OmpⅡ respectively,were successfully obtained and the recombinant expression vector(pGEX-2T-Vibr-Aero-his) was combined by fusion PCR.Expression of the bivalent Omp was induced over night by 1.0 mmol/L IPTG at 16 ℃ when the concentration of 600 nm optical density of the bacteria was 0.6-1.0,and the expressed protein with the molecular weight of 82.2 ku was obtained and purified by HisPur Ni-NTA Resin and Kits.Eels were immunized with the bivalent OMP after purified and renatured by dialysis to determine the antibody titer.Compared with eels in PBS group,the serum titers of anti-V.vulnificus and A.hydrophila antibody in eels of Omp group,showed significant(P<0.05) increase on 14 d and 21 d,and the different between the two groups reached very significant levels(P<0.01) on 28 d and 42 d.The study laid a foundation for the vaccine research and applicationin aquaculture of the bivalent outer membrane protein.

相似文献/References:

[1]张芳芳,熊静,段明珠,等.日本鳗鲡COX-2基因的克隆、鉴定及表达分析[J].集美大学学报(自然版),2015,20(5):321.
 ZHANG Fang-fang,XIONG Jing,DUAN Ming-zhu,et al.Molecular Cloning,Characterization and Expression of COX-2 in Japanese eel,Anguilla japonica[J].Journal of Jimei University,2015,20(3):321.

更新日期/Last Update: 2017-09-10