Immunogenicity analysis of a recombinant protein CPSIT_p7 and its expression in HeLa cells during persistent Chlamydophila psittaci infection
10.3760/cma.j.issn.0254-5101.2014.08.007
- VernacularTitle:CPSIT_p7重组表达产物免疫原性检测及其在鹦鹉热嗜衣原体持续性感染中表达的研究
- Author:
Qingzhi HE
;
Huaicai ZENG
;
Zhiqing LI
;
Chuan WANG
;
Yanqun HU
;
Zhixi CHEN
;
Yimou WU
- Publication Type:Journal Article
- Keywords:
Chlamydophila psittaci;
CPSIT_p7;
Cloning and expression;
Immunogenicity;
Per-sistent infection
- From:
Chinese Journal of Microbiology and Immunology
2014;(8):604-608
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct a prokaryotic expression plasmid for CPSIT_p7 gene from Chlamydophila psittaci ( Cps) 6BC strain and to evaluate immunogenicity of the recombinant protein His-CPSIT_p7 and detect its dynamic expression at mRNA and protein levels in HeLa cells during persistent Cps infection.Methods The fusion protein His-CPSIT_p7 was expressed in E.coli BL21 and purified by Ni-NTA affinity chromatography .BALB/c mice were immunized with the recombinant protein to prepare polyclonal antibody for evaluation of the immunogenicity of His-CPSIT_p7 by ELISA.Penicillin sodium was used to establish a model of Cps persistence infection .RT-PCR and Western blot assay were performed to de-tect the expression of CPSIT_p7 at mRNA and protein levels during Cps persistent infection .Results The fusion protein His-CPSIT_p7 was successfully expressed with the use of constructed recombinant expression plasmid pET30 a-CPSIT_p7 and purified .ELISA result showed that the specific antibody titer against CPSIT_p7 reached 1 ∶1 000 000 on the 40th days after immunization .The expression of CPSIT_p7 at mRNA and protein levels were increased in a time-dependent manner in Cps-infected HeLa cells .The peak of mRNA level was reached at the time point of 36 hours after infection , followed by a time-dependent decrease during Cps acute infection .However , the expression of CPSIT_p7 at mRNA and protein levels were not decreased until 60 hours after infection during Cps persistent infection .Conclusion His-CPSIT_p7 protein was suc-cessfully expressed in the prokaryotic expression system and purified , showing an advantage of good immuno-genicity.Highly expressed CPSIT_p7 at mRNA and protein levels were detected during Cps persistent infection.
