Objective:To clone the rabies virus G protein(RVG) gene fragments into prokaryotic expressing vector, and to express and purify RVG, which could be applied as diagnostic antigen for indirect ELlSA assay to detect rabies virus antibodies. Methods:The G gene of rabies virus was amplified by RT-PCR with a pair of specific primers designed according to the relevant sequence of GenBank. The PCR product was cloned into prokaryotic expression vector pGEX-6P-1 to construct the expression plasmid pGEX-RVG. The positive recombinant plasmid was transformed into host bacteria E.coli BL21(DE3) and induced with IPTG. In order to determine the optimal induction condition for soluble expression, optimization condition was studied. The glutathione S-transferase (GST) affinity chromatography was used to purify the recombinant protein, and the biological activity of RVG was analyzed by Western blot. Results:The prokaryotic expression vector pGEX-RVG were successfully constructed. After inducted by IPTG, the fusion protein about 36 000 was observed,which was the same as expected. The recombinant fusion protein has a fine biological activity which was confirmed by Western blot. Conclusion:The rabies virus G protein was successfully expressed and purified, which could be provided diagnostic antigen for indirect ELlSA assay for the detection of rabies virus antibodies.