Objective To construct the eukaryotic expression vector PCI-dhfr-λ1 and PCI-dhfrSP163-λ1 which linked the enhancer SP163 with interferon λ1.Then express the interferon λ1 in CHO (dhfr-) cells.Methods Using PCR method to introduce the restriction enzyme sites and through the fusion PCR binding the enhancer with the interferon λ1.After sequenced,λ1 and SP163-λ1 was inserted into PCI-dhfr forming the expression vector PCI-dhfr-λ1 and PCI-dhfr-SP163-λ1 which was constructed successfully confirming by sequencing.Then the expressing vectors were transfected into CHO(dhfr-) cells using liposome transfection method and interferon λ1 protein was assayed with indirect immunofluorescence and Western Blot.Using cytopathic effect inhibition evaluated the antiviral activity of interferon λ1.Results Successfully constructing the eukaryotic expression vectors of interferon λ1 and the vectors could express interferon λ1.The result of immunofluorescence showed the enhancer developed the expression of interferon λ1.Detecting the interferon λ1 in CHO (dhfr-) cells after transfecting 48 hour using Western Blot.The cytopathic effect inhibition showed the expressed interferon λ1 has the antiviral activity.Conclusion Successfully expressed the interferon λ1 in CHO(dhfr-) cells and the protein possesses antiviral activity,which may supply a valuable basis for building the stable cell line of interferon λ1.