中国农业科学院农业信息研究所 中国农学会
Agricultural Information Institute of CAAS China Association of Agricultural Science Societies
En
Ch
中国农业科学院农业信息研究所 中国农学会 主办

AI检索

订阅

返回顶部
当前位置: 首页 > 文章 > 流产衣原体CPAF基因的克隆序列分析及原核表达 华北农学报 2018 (3) 86-91
Position: Home > Articles > Cloning,Sequence Analysis and Prokaryotic Expression of CPAF Gene of Chlamydia abortus Acta Agriculturae Boreali-Sinica 2018 (3) 86-91

流产衣原体CPAF基因的克隆序列分析及原核表达

作  者：
刘萍;马晓霞;周小凯;马鹏;常秋燕;李林杰;李凌浩;马忠仁
关键词：
流产衣原体;CPAF;克隆;序列分析;原核表达
摘  要：
为了研究流产衣原体蛋白酶样活性因子(CPAF)的免疫原性,建立快速有效的流产衣原体抗体检测方法,根据Gen Bank中公布的流产衣原体基因组序列,设计并合成了流产衣原体CPAF蛋白编码基因的特异性引物,以流产衣原体标准菌株基因组为模板,经PCR扩增获得长度为1 809 bp目的基因片段;对获得的目的基因进行测序,并用生物信息学软件进行预测分析;再将该片段插入原核表达载体p ET-30a中,经双酶切、测序鉴定;构建成功的表达质粒转化大肠杆菌BL21(DE3),并用IPTG对目的蛋白进行诱导表达,通过对诱导条件的优化,确定诱导表达的最佳条件,表达产物用SDS-PAGE和Western Blot检测。结果显示,成功克隆流产衣原体CPAF蛋白的编码基因,该编码产物可能是定位于宿主细胞质中的一种衣原体分泌性蛋白酶,分子内具有多个可能的抗原表位;重组表达质粒经IPTG诱导后,表达分子质量为67.6 ku的CPAF蛋白,该重组蛋白能与抗His-tag单克隆抗体以及流产衣原体阳性血清特异性结合,具有良好的反应原性。结果表明,以大肠杆菌表达系统重组表达的流产衣原体CPAF蛋白具有良好的免疫原性,可作为动物流产衣原体病的血清学诊断及亚单位疫苗研究的候选抗原。
译  名：
Cloning,Sequence Analysis and Prokaryotic Expression of CPAF Gene of Chlamydia abortus
作  者：
LIU Ping;MA Xiaoxia;ZHOU Xiaokai;MA Peng;CHANG Qiuyan;LI Linjie;LI Linghao;MA Zhongren;Key Laboratory of Bioengineering and Biotechnology of State Ethnic Affairs Commission,Northwest Minzu University;Gansu Engineering & Technology Research Center for Animal Cell;Life Science and Engineering College,Northwest Minzu University;
单  位：
LIU Ping%MA Xiaoxia%ZHOU Xiaokai%MA Peng%CHANG Qiuyan%LI Linjie%LI Linghao%MA Zhongren%Key Laboratory of Bioengineering and Biotechnology of State Ethnic Affairs Commission,Northwest Minzu University%Gansu Engineering & Technology Research Center for Animal Cell%Life Science and Engineering College,Northwest Minzu University
关键词：
Chlamydia abortus;;CPAF;;Clone;;Sequence analysis;;Prokaryotic expression
摘  要：
In order to investigate the immunogenicity of Chlamydial protease like activity factor( CPAF) of C. abortus and establish a rapid method for detection of Chlamydia abortus( C. abortus),a pair of specific primers for CPAF of C. abortus were designed and synthesized. The gene was amplified by PCR from C. abortus reference strain. The obtained gene sequence was sequenced and analyzed by bioinformatics software. The obtained gene of 1 809 bp was inserted into plasmid p ET-30 a. The recombinant plasmid was identified by digesting and sequencing. Then the successfully constructed recombinant plasmid was transformed into E. coli BL21( DE3) strain as host cell for expression of the recombinant protein by IPTG induction. The production was detected by SDS-PAGE and Western Blot analysis. The CPAF gene of Chlamydia abortus was successfully cloned and analyzed. This gene was likely to be located in the cytoplasm of host cell,playing a role as a proteolytic enzyme with several potential antigenic epitope. As shown by SDS-PAGE and Western Blot analysis,a 67. 6 ku recombinant protein was expressed by IPTG induction,which could combine with anti-His-tag monoclonal antibody and C. abortus positive serum with well reactiongenicity. These results suggested that the recombinant CPAF could be used as a potential antigen for serological tests and vaccine development of C. abortus.

相似文章

导出引用
计量
文章访问数: 9
HTML全文浏览量: 0
PDF下载量: 0

所属期刊

推荐期刊

友情链接Friendly link
农业信息研究所Agricultural information institute 农业智能知识服务平台Agricultural intelligent knowledge service platform 农业学术服务平台Agricultural Knowledge Service System 中国知网CNKI 维普网VIP 万方WANFANG DATA
联系我们Contact us

电话Tell：010-82109652-808

邮箱E-mail：agrijournal@caas.cn

服务Service
关于我们About us 意见建议Opinions and suggest
版权说明Copyright
单位：中国农业科学院农业信息研究所 中国农学会 © 2017中国农业科学院农业信息研究所 ICP备案号：京ICP备09089781号-46