猪圆环病毒2型Cap蛋白家蚕表达体系的构建与鉴定

Construction and Identification of Bombyx mori Expression System for Porcine Circovirus Type 2 Cap Protein

为了用家蚕杆状病毒表达系统表达猪圆环病毒2型(PCV2)Cap蛋白,将不同株系的PCV2 Cap蛋白基因、人工改造的Cap蛋白突变体基因克隆到家蚕杆状病毒转移载体,用共转染技术成功构建了重组杆状病毒。将重组杆状病毒感染家蚕,成功获得了大量Cap蛋白的表达产物。结果表明,纯化蛋白主要为病毒空衣壳粒子,推算PCV2空衣壳在家蚕中的表达量可达1.2 mg/蛹~1.6 mg/蛹。电镜观察可见到完整的空衣壳粒子,胶体金标记免疫电镜观察可见到完整的胶体金粒子,金粒大小为10 nm。

In order to express PCV2 Cap protein using silkworm baculovirus expression system,different strains of PCV2 Cap protein gene and artificially modified Cap protein mutant gene were cloned into Bombyx mori baculovirus transfer vector.The recombinant baculovirus for expression was successfully constructed by co-transfection technology.A large number of Cap protein expression products were successfully obtained by recombinant baculovirus infection.The identification results showed that the purified protein was mainly viral empty capsid particles,and the expression level of PCV2 empty capsid was estimated to be 1.2-1.6 mg/pupa.Complete empty capsid particles were observed by electron microscopy,and complete colloidal gold particles were observed by colloidal gold labeled immunoelectron microscopy.The size of gold particles was 10 nm.