title: Prokaryotic expression and immunogenicity of 56-kDa protein of Orientia tsutsugamushi strain Karp.
abstract: To express the 56-kDa protein of O. tsutsugamushi strain Karp, this protein gene was cloned into pET30a(+) before transforming into host bacteria, E. coli Rossetta. Specificity of the recombinant protein was assessed by ELISA using rabbit sera against common members of the order Rickettsiae and 10 other pathogenic bacteria. After IPTG induction, SDS-PAGE analysis of isolated protein demonstrated a band at approximately 46-kDa. Western blot and mass spectrometry analysis proved that the recombinant protein was expressed successfully. Specificity analysis demonstrated that all sera were negative, except sera against O. tsutsugamushi strains TA763, TH1817 and Kato, B. quintana, A. phagocytophilum, E. chaffeensis and B. bacilliformis. The purified protein was used to immunize BALB/c mice and polyclonal antisera were harvested. By examination of IFA and ELISA, the highest titer of the polyclonal antibodies reaches 1:1600. The recombinant 56-kDa protein in the study is valuable for developing a simple and rapid diagnostic test and vaccine for O. tsutsugamushi.
authors.affiliation: Department of Rickettsiology, National Institute for Communicable Disease Control and Prevention, China CDC, Beijing, China.
authors.forename: Yuan-Yuan
authors.initials: YY
authors.lastname: Wang
authors.forename: Qiang
authors.initials: Q
authors.lastname: Chen
authors.forename: Qiang
authors.initials: Q
authors.lastname: Yu
authors.forename: Li-Juan
authors.initials: LJ
authors.lastname: Zhang
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D003429:Cross Reactions; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology; D015870:Gene Expression; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D012285:Orientia tsutsugamushi / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D012612:Scrub Typhus / Q000276:immunology / Q000382:microbiology
chemical_list: D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D011994:Recombinant Proteins