Expression And Purification Of Structure Protein Of SARS-CoV And Serological Evidence Of SARS-CoV Like Viruses Spread Widely In Domestic Palm Civets In China

This thesis includes four chapters. In chapter one, a brief introduction of SARS-CoV and recent advances in SARS-CoV research were presented. The background and the aim of the thesis were also described. Chapter two is a report on prokaryotic expression of structure Protein genes from SARS-CoV and generation of specific antibodies. Eight structure Protein genes (S, S₁, S₂, S_â–³,M, E, N,ORF03,ORF08)were amplified by RT-PCR from the throat swabs collected from SARS patients. The PCR fragments were cloned into expression vector pGEX-KG plasmids. After the induction of IPTG, the E. coli BL21 which containing S1_, S₂, S_â–³,M,E, N,ORF08 genes expressed proteins with molecular weights of 98 kDa,86 kDa, 58 kDa,52 kDa,34 kDa,74 kDa,38 kDa, respectively, which were in agreement with the expectation. The purified expressed proteins were used to immune rabbits separately. Western Blot analysis using the antibodies derived from the rabbits indicated that these antibodies could react specifically with the target proteins and were suitable to be used for further functional analysis and detection of SARS. And we detected immunogenic of recombinant protein, Inactivated SARS-CoV was used as an antigen to detect anti-SARS-CoV antibodies, result showed that recombinant nucleocapsid protein of SARS-CoV showed good immunogenic activity. In chapter three, 110 serum specimens were collected from different civet farms in China in 2003 and 2004, we examined these specimens using different methods. The result showed that 84% of the domestic palm civets have antibody to SARS-CoV, serum specimens from different area contained different levels of antibody to SARS-CoV, which suggests SARS-CoV-like virus widespread among domestic civets on farms in China. In chapter four,It is discussion and conclusion of the research.