关键词: 猪链球菌2型, 溶菌酶释放蛋白, 胞外蛋白因因子, 融合表达, 中和表位

Abstract: Two fragments of mrp gene (1 801-2 513bp) and epf gene (1 783-2 563bp ) were amplified from genomic DNA of Streptococcus suis type 2 isolate strain SS2-1 by polymerase chain reaction (PCR) technique. The PCR products were later cloned into plasmid vector pET-32a via restriction endonuclease and then transformed into host strain BL21. Through relevant endonuclease digest , the positive recombinant bacteria were identified, which were expressed by IPTG of optimal concentration, respectively. Also, the antigenicity of expressed recombinant proteins (rMRP and rEPF) were determined by Western-blot. After verified positive immunity, the fragments of mrp and epf linked by PCR was cloned into pET-32a again. By the above method with IPTG, the recombinant protein consisted of rMRP and rEPF with molecular weight 74kD was obtained. Next, the antigenicity analysis by Western-blot showed that the recombinant protein had the conserved epitopes of MRP and EPF, respectively. In order to further determine the antigenicity of recombinant proteins (rMRP, rEPF and rMRP-EPF), rabbits were immunized, positive (immunity with bacterial vaccine) and negative controls (non-immunity) were set. After 21d, challenged by Streptococcus suis type 2 isolate strain SS2-1,rMRP-EPF immunized rabbits were protected more than alone. Compared to rMRP 25%,rEPF 0%, bacterial vaccine 100%, rMRP-EPF beared protection rates of 50%. It is concluded that rMRP-EPF possesses good antigenicity.

Key words: Streptococcus suis type 2, Muramidase-released protein, Extracellular protein factor, Fusion expression, Epitope