关键词: 禽流感, H5亚型, 鸡β－actin启动子, DNA疫苗, 免疫保护

Abstract: 【Objective】 To improve the protective efficacy of H5 subtype avian influenza DNA vaccine, the HA gene of A/Goose/GuangDong/1/96(H5N1) [GD/1/96(H5N1)] was inserted into pCAGGS, which contains chickenβ-actin promoter, and was designated as pCAGGHA5. 【Method】The monolayer 293T cells were transfected by pCAGGHA5 and pCIHA5, which contained HA gene of GD/1/96(H5N1). The HA protein expression was detected by indirect immunofluorescence assay 24-48 h after transfection and confirmed by western blotting analysis. To evaluate the vaccine efficacy, groups of 3-week-old SPF chickens were intramuscular inoculated with 100 μg or 10 μg of pCIHA5, pCAGGHA5, respectively. A group of chickens were injected with 200 μl PBS as control. Four-weeks after the single vaccination, all chickens were challenged with 100 LD50 of highly pathogenic GD/1/96(H5N1). 【Results】 In vitro expression results indicated the gene in the pCAGGS vector was better expressed than that in the pCI vector. The chickens in the 100μg and 10μg pCAGGHA5 immunized groups were completely protected from virus challenge (no disease signs, no virus shedding and no deaths), and 100μg pCIHA5 immunized groups only provided partial protection, while the chickens in the pCIHA5 immunized group and the control group died within ten days. 【Conclusion】 Results shown that expressive vector optimization enhanced the protective efficacy of H5 subtype avian influenza DNA vaccine.

Key words: Avian influenza, H5 subtype, Chickenβ-actin promoter, DNA vaccine, Protective efficacy