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

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