Cloning,Sequence Analysis and Prokaryotic Expression of CPAF Gene of Chlamydia abortus
LIU Ping1,2,3, MA Xiaoxia1,2,3, ZHOU Xiaokai1,2,3, MA Peng1,2,3, CHANG Qiuyan1,2,3, LI Linjie1,2,3, LI Linghao1,2,3, MA Zhongren1,2,3
1. Key Laboratory of Bioengineering and Biotechnology of State Ethnic Affairs Commission, Northwest Minzu University, Lanzhou 730030, China;
2. Gansu Engineering & Technology Research Center for Animal Cell, Lanzhou 730030, China;
3. Life Science and Engineering College, Northwest Minzu University, Lanzhou 730030, China
Abstract：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 pET-30a. 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.
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2018, Vol. 33 
