关键词: 免疫活性

Abstract:

【Abstract】 Objective To clone and express the Tp0136 gene of Treponema pallidum, to purify the recombinant protein and to evaluate its immunocompetence. Methods The full-length Tp0136 gene was synthesized, then subcloned into the expression vector pET28a to construct a recombinant plasmid, pET28a-Tp0136, which was subsequently transfected into E.coli Rosetta for protein expression. The recombinant protein was purified with nickel-nitrilotriacetic acid （Ni-NTA） affinity chromatography, and identified by using sodium dodecyl sulfate polyacrylamide gel electropheresis（SDS-PAGE） and Western blot. New Zealand rabbits were immunized with the recombinant Tp0136 （rTp0136）protein, and anti-Tp0136 polyclonal antibodies in sera of the rabbits were examined by indirect enzyme linked immunosorbent assay （ELISA） with rTp0136 protein as the coating antigen. Also, positive sera were obtained from patients with syphilis and examined by Western blot to identify the immunoreactivity of the rTp0136 protein. Results The E.coli expression vector pET28a-Tp0136 was constructed successfully, and rTp0136 protein was also successfully expressed with a molecular weight of about 50 kD and a purity above 95%. High titres of anti-rTp0136 antibodies were detected in sera of rabbits immunized with the rTp0136 protein, and Western blot showed that the rTp0136 could specifically react with the sera from syphilitic patients, which proved the high immunogenicity and immunoreactivity of the recombinant protein. Conclusions The full-length Tp0136 membrane protein is successfully expressed with a high immunocompetence, and Tp0136 membrane protein may play an important role in the pathogenic mechanisms of Treponema pallidum.

Key words: Immunocompetence