BmCPV多角体蛋白对异源蛋白的包埋

为了解家蚕质型多角体病毒(BmCPV)的多角体蛋白对异源蛋白的包埋作用,将BmCPV的多角体蛋白基因克隆至昆虫细胞表达载体pIZT/V5-His,转染家蚕卵巢培养细胞(BmN)后通过吉欧霉素(zeocin)筛选获得稳定表达多角体蛋白的细胞株。倒置荧光显微镜观察发现,转化BmN细胞有轻度的病理变化,细胞质中有呈绿色荧光的多角体蛋白结晶,但从转化BmN细胞中纯化的多角体无明显的绿色荧光。以修饰型线性化家蚕杆状病毒BmPAK6感染稳定转化BmN细胞,96 h后纯化多角体,回复感染显示多角体裂解液能感染家蚕BmN细胞,表明BmCPV多角体蛋白能包埋BmPAK6。纯化的多角体及多角体裂解液均能用X-gal显色,表明BmPAK6表达的β-半乳糖苷酶也能被BmCPV的多角体蛋白包埋。研究结果可为获得具有多角体的重组杆状病毒以及开发利用质型多角体蛋白包埋异源蛋白的功能提供新的技术途径。

Immobilization of Foreign Protein by Polyhedra of BmCPV

In order to study the function of Bombyx mori cytoplasmic polyhedrosis virus(BmCPV) polyhedra incorporating heterologous proteins,the BmCPV polyhedrin gene was cloned into insect-expressed vector pIZT/V5-His and then transformed into BmN cells.The cell line stably expressing polyhedrin was obtained by screening with antibiotic zeocin.Using the inverted fluorescence microscope,we found that the transformed cells had a little pathological change,and in the cytoplasm there were some green fluorescent polyhedrin crystallizations,but the polyhedra purified from the transformed cells did not show obvious green fluorescence.The stably transformed BmN cells were infected with the modified linear baculovirus BmPAK6 and the polyhedra were purified 96 h later.Artificial re-infection showed that the lysate of polyhedra could infect BmN cells,indicating that the polyhedrin of BmCPV could embed BmPAK6.Both the purified polyhedra and its lysis supernate could be coloured by X-gal,indicating that the β-galactosidase expressed by BmPAK6 could also be incorporated into the polyhedra.These results provide novel technical references to obtaining recombinant baculovirus with polyhedra as well as exploiting and utilizing the capability of cytoplasmic polyhedrin to embed heterologous proteins.