[Objective] In this study, a new chimeric protein SEG expressed in previous work was applied to evaluate its translocating efficiency of shRNA to rabies virus infected cells in mice, meanwhile, the capability of anti-rabies virus was investigated.[Methods] Rabies virus strain CVS-24 was inoculated into the hind leg to establish a mouse model of rabies in a dose of 50 LD₅₀; 12 h thereafter the mice were injected intravenously with shRNA-producing plasmid mixed with SEG. To test shRNA delivery, single-cell suspensions from brain, spleen and liver were examined by flow cytometry. Rabies virus in brain tissue of mice was detected by qRT-PCR, RT-PCR, western blot and directed immunofluorescence assay. Mice were monitored for survival and serum samples were tested for IFN-α levels.[Results] No green fluorescent protein(GFP) was seen in the spleen or liver, suggesting that SEG allows specific targeting of RV-infected cells. RT-PCR and western blot showed that mice treated with SEG-shRNA had lower rabies virus RNA and protein levels than the controls. Real-time PCR showed that rabies virus was reduced 4.88 fold compared to the mock cells. Survival of RV-infected mouse showed a significant protection from rabies virus infection by SEG-shRNA treatment. The survival was up to 50% whereas the control group all died. IFN was not induced in SEG-shRNA treated animals.[Conclusion] shRNA-producing plasmid was specifically delivered into rabies virus infected cells using the SEG protein, and effectively inhibited rabies virus gene expression and replication in vivo. SEG-shRNA can be used for adjuvant treatment for rabies.