1.Cloning,Expression and Immunization of The Hypoxanthine-guanine Phosphoribosyltransferase for Schistosoma japonicum Chinese Strain
Junlong YU ; Shiping WANG ; Zhuo HE ; Gan DAI ; Wenkai LI ; Xiaoxin JIANG ; Shaohua ZENG ; Xiaoqin XIAO ; Shaorui XU ; Zhiyue L ; Xianchu PENG ; Songhua ZHOU ; Xueqin LIU
Progress in Biochemistry and Biophysics 2006;0(07):-
A 1 270 bp full-length cDNA fragment was obtained from the Schistosoma japonicum (Chinese strain) adult cDNA library after the 3′ and 5′ ends of the incomplete expression sequence tag (EST) of hypoxanthine-guanine phosphoribosyltransferase of Schistosoma japonicum (SjHGPRT) were amplified by the anchored PCR with 2 pairs of primer that were designed according to the published incomplete SjHGPRT EST and the sequence of multiclone sites of library ?gt11 vector. Sequence analysis indicated that this fragment, with an identity of 82% to hypoxanthine-guanine phosphoribosyltransferase of Schistosoma mansoni (SmHGPRT), contained a complete open reading frame(ORF). The deduced amino acid sequence showed 83% identity to that of SmHGPRT. This fragment was cloned into the prokaryotic expression vector pQE30, and subsequently sequenced and expressed in Escherichia coli. SDS-PAGE revealed that M of the recombinant protein was about 28 ku. Western-blot analysis showed that the recombinant protein was recognized by the polyclonal antisera from rabbits immunized with Schistosoma japonicum adult worm antigen. Mice vaccinated with recombinant protein revealed significant worm burden, liver eggs per gram (LEPG), fecal eggs per gram (FEPG) and intrauterine eggs of the female worms reduction percentage, compared with the controls. Taken together, the SjHGPRT full-length cDNA can be cloned and expressed in E.coli as a recombinant protein that elicited immunity against the challenge infection with Schistosoma japonicum, indicating its potential as a partial protection vaccine candidate.
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