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Cloning,expression and identification of Toll like receptor interacting protein gene of Schistosoma japonicum

Objective To clone and express Schistosoma japonicum Toll like receptor interacting protein(SjTollip) in prokary-otic expression system and analyze its stage specific transcription and expression.Methods The encoding sequence selected from S.japonicum cDNA library was amplified by PCR.The SjTollip gene obtained was subcloned into pET 28a,then transformed into E.coli BL21 and induced with IPTG for expression.The expressed protein was purified with Ni NTA resin.Total RNA were extracted from different stages of S.japonicum.The immune rabbit sera were prepared by immunizing New Zealand white rabbits with purified recombinant SjTollip protein.RT PCR and Western blotting were used to analyze the transcription and expression level at the different stages and the immunogenicity.Results The expression vector of SjTollip/pET 28a was constructed and expressed as inclusion bodies(M r 24 000).The recombinant protein rSjTollip was specifically recognized by the S.japonicum infected rabbit serum.SjTollip showed lower transcription level in stages including cercariae and male worms.Western blotting analysis showed that the target protein was detected in all stages.The expression level in stages including schistosomulum and female worm was much higher.Conclusions The SjTollip transcription and expression level at the different stages of S.japonicum is different,and this gene might be a potential candidate for target of vaccine,drug and diagnosis.