Abstract: IFN-λ as a pleiotropic cytokine belongs to type-Ⅲ interferons and demonstrate fundamental role against viruses that escape from the host immune system. In order to efficiently express Ovis aries λ3 interferon (OvIFN-λ3) with high antiviral activity by using Bombyx mori baculovirus expression system, firstly OvIFN-λ3 gene was optimized and synthesized, then cloned into transfer vector pVL1393 to obtain recombinant plasmid pVL1393-OvIFNλ3, after that, the recombinant plasmid and inactivated-rescued Bombyx mori baculovirus shuttle vector reBmBac DNA were co-transfected into Bombyx mori ovarial passage cell Bm-N to obtain recombinant baculovirus containing OvIFN-λ3 gene. The recombinant baculovirus was then infected with the 5th instar silkworm larvae and the hemolymph of the silkworms was collected after the onset of the disease. The anti-virus experiment of recombinant vesicular stomatitis virus expressing green fluorescent protein (VSV-GFP) was carried out on sheep kidney cells by micro cytopathic inhibition method, and initial antiviral activity of OvIFN-λ3 was determined to be (6.5±0.27)×105 U/mL. And plaque assay was performed to purify the recombinant baculovirus and surprisingly it was demonstrated that antiviral potential of purified recombinant virus was (3.1±0.42)×106 U/mL, it indicated that the expression level was increased. The silkworm-baculovirus expression system provided a new method for mass-scale production and application of Ovis aries λ3 interferon products.

Key words: Ovis aries λ3 interferon, silkworm-baculovirus expression system, antiviral activity assay

摘要： λ干扰素作为一种多效性细胞因子属于Ⅲ型干扰素，研究表明其能够对Ⅰ型和Ⅱ型干扰素逃逸的病毒起作用。为了利用家蚕杆状病毒表达系统高效表达出具有高抗病毒活性的羊λ3干扰素（OvIFN-λ3），首先将OvIFN-λ3基因优化后合成，将其克隆至转移载体pVL1393中，得到重组质粒pVL1393-OvIFNλ3，再将重组质粒与失活拯救型家蚕杆状病毒穿梭载体reBmBac DNA共转染家蚕卵巢传代细胞Bm-N，得到含OvIFN-λ3基因的重组杆状病毒。随后利用重组杆状病毒感染五龄起蚕，待其发病后收集蚕血淋巴。采用微量细胞病变抑制法在羊肾细胞上进行重组表达绿色荧光蛋白的水疱型口炎病毒（recombinant vesicular stomatitis virus expressing green fluorescent protein，VSV-GFP）的攻毒实验，测定OvIFN-λ3抗病毒活性可达（6.5±0.27）×105 U/mL；利用空斑筛选法筛选重组病毒，感染家蚕后，测定其效价最高可达（3.1±0.42）×106 U/mL，表达量明显提高。家蚕杆状病毒表达系统为羊λ3干扰素产品的大量生产应用提供了新方法。

关键词: 羊&lambda, 3干扰素, 家蚕杆状病毒表达系统, 抗病毒活性检测