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Subcloning and expression of gene encoding the cystein proteinase of Rhipicephalus haemaphysaloides tick and the immune protective analysis of its expressed products



Subcloning and expression of gene encoding the cystein proteinase of Rhipicephalus haemaphysaloides tick and the immune protective analysis of its expressed products



Chinese Journal of Zoonoses ober; 21(10): 871-874



To subclone and express the gene encoding the cystein proteinase of Rhipicephalus haemaphysaloides tick and to analyze the immune protection of its expressed products, the cDNA of cys A and cysB genes were cloned into vector pET-32a, then transformed to E . coli BL21 cells and expressed by the induction with IPTG. The expressed products were purified and used as antigen to immunize rabbits in order to evaluate its immune protective effect. in which the rabbits in the vaccinated group were immunized with 300[mu]g at first and 150 [mu]g in the next two injections of the recombinant protein emulsified with Freund's complete adjuvant (FCA), while those in the control group were injected with PBS emulsified with FCA, and the whole course of immunization consisted of triple injections at weeks 0,2 and 4 respectively. The rabbits were challenged with R. haemaphysaloides ticks 2 weeks after the final injection, and the effect of immune protection was observed after challenging. It was found that the fusion protein Trx-CysA and Trx-CysB were highly expressed in E E. coli in insoluble form and the inclusion bodies after induction by IPTG. The reduction rates of tick attachment in the vaccinated groups with both Trx-CysA and Trx-CysB were 17% ; while the reduction in tick engorgement in the vaccinated groups with Trx-CysA and Trx-CysB were 42% and 22 % respectively. It is evident from the above observations that the cysA and cysB genes can be highly expressed in E . coli after subcloning into vector pET-32a, and the fusion proteins Trx-CysA and Trx-CysB can significantly induce a certain degree of the anti-tick protective immunity.

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