以CotB作为融合基序在枯草杆菌芽孢表面展示鸡白痢沙门氏菌OmpC的研究
详细信息 查看官网全文
- 英文论文题名：Display of OmpC of Salmonella pullorum on Bacillus subtilis spores using CotB as a fusion partner
- 论文作者：刘明刚 ; 刘红露 ; 戴茜茜 ; 徐毓琴 ; 张飞燕 ; 谷笑笑 ; 潘康成
- 英文论文作者：LIU Ming-gang ; LIU Hong-lu ; DAI Xi-xi ; XU Yu-qin ; ZHANG Fei-yan ; GU Xiao-xiao ; PAN Kang-cheng ; Animal Microecology Institute ; College of Veterinary Medicine ; Sichuan Agricultural University ; Key Laboratory of Animal Diseases and Human Health of Sichuan Province
- 年：2016
- 作者机构：四川农业大学动物医学院动物微生态研究中心;动物疫病与人类健康四川省重点实验室;
- 论文关键词：沙门氏菌 ; 枯草芽孢杆菌 ; 芽孢表面展示 ; 粘膜疫苗
- 英文论文关键词：Salmonella ; Bacillu subtilis ; spore surface display ; mucosal vaccine
- 会议召开时间：2016-11-04
- 会议录名称：中国畜牧兽医学会动物微生态学分会第五届第十二次全国学术研讨会论文集
- 语种：中文
- 分类号：S852.61
- 学会代码：ZGXJ
- 会议名称：中国畜牧兽医学会动物微生态学分会第五届第十二次全国学术研讨会
- 会议地点：中国山东青岛
- 主办单位：中国畜牧兽医学会动物微生态学分会
- 学会名称：中国畜牧兽医学会
- 页数：1
- 文件大小：77k
- 原文格式：O
- 会议级别：全国
摘要
本文旨在通过使用枯草芽孢杆菌芽孢衣壳蛋白CotB作为融合基序,构建表面展示鸡白痢沙门氏菌外膜蛋白OmpC的重组芽孢,为防控沙门氏菌感染提供新的思路。分别PCR扩增鸡白痢沙门氏菌OmpC编码基因与含自身启动子的芽孢衣壳蛋白CotB编码基因,以质粒pDG364为基础进行二者融合并构建整合型重组质粒pDG364-cotB-ompC,将融合基因整合于枯草芽孢杆菌168 amyE基因位点,通过淀粉酶活性、PCR技术、免疫荧光显微镜进行分析。以芽孢衣壳蛋白CotB为融合基序,成功构建获得携带CotB和OmpC融合基因的整合型重组质粒pDG364-cotB-ompC,并经同源双交叉重组将其整合于枯草芽孢杆菌168基因组上,通过淀粉酶活性分析与PCR鉴定表明该融合基因成功整合于amyE基因位点,使用OmpC特异性抗体探测的免疫荧光显微镜分析检测到特异性的荧光信号。结果表明,鸡白痢沙门氏菌OmpC以具有生物活性的形式成功表达并展示于枯草芽孢杆菌168芽孢表面。
The aim of this study was to construct recombinant Bacillus subtilis spores expressing the OmpC antigen fused to the spore coat protein CotB and develop a new way for prevention and control of Salmonella infections.The cotB with its promoter and ompC genes were amplified by PCR from the genomic DNA of Salmonella pullorum and Bacillus subtilis,respectively.The cotB-ompC gene fusion was obtained by cloning ompC in frame of cotB in the integrative vector pDG364,yielding plasmid pDG364-cotB-ompC.The obtained fusion gene was integrated into the B.subtilis168 chromosome at the amyE locus by a double crossover event.Several clones were sequentially tested by PCR,amylase activity assay and immunofluorescence microscopy.The results indicated that the correct insertion of cotB-ompC gene fusion at amyE locus on B.subtilis168 chromosome and OmpC was successfully expressed and displayed on spores surface with biological activity.
The aim of this study was to construct recombinant Bacillus subtilis spores expressing the OmpC antigen fused to the spore coat protein CotB and develop a new way for prevention and control of Salmonella infections.The cotB with its promoter and ompC genes were amplified by PCR from the genomic DNA of Salmonella pullorum and Bacillus subtilis,respectively.The cotB-ompC gene fusion was obtained by cloning ompC in frame of cotB in the integrative vector pDG364,yielding plasmid pDG364-cotB-ompC.The obtained fusion gene was integrated into the B.subtilis168 chromosome at the amyE locus by a double crossover event.Several clones were sequentially tested by PCR,amylase activity assay and immunofluorescence microscopy.The results indicated that the correct insertion of cotB-ompC gene fusion at amyE locus on B.subtilis168 chromosome and OmpC was successfully expressed and displayed on spores surface with biological activity.
引文
[1]LEVINE M M.Enteric infections and the vaccines to counter them:future directions[J].Vaccine,2006,24(18):3865-3873.
[2]CLEMENS J.Evaluation of vaccines against enteric infections:a clinical and public health research agenda for developing countries[J].Philos.T.R.Soc.B,Biological sciences,2011,366(1579):2799-2805.
[2]CLEMENS J.Evaluation of vaccines against enteric infections:a clinical and public health research agenda for developing countries[J].Philos.T.R.Soc.B,Biological sciences,2011,366(1579):2799-2805.