摘要
In the late phase of Bombyx mori nucleopolyhedrovirus(BmNPV)infection,a large amount of polyhedra appear in the infected cell nucleolus,these polyhedra being dense protein crystals protecting the incorporated virions from the harsh environment.To investigate whether the foreign protein could be immobilized into the polyhedra of BmNPV,two recombinant baculoviruses were generated by a novel BmNPV polyhedrin-plus(polh +)Bac-to-Bac system,designated as vBmBac(polh +)-enhanced green fluorescent protein(EGFP)and vBmBac(polh +)-LacZ,which can express the polyhedrin and foreign protein simultaneously.Light microscopy analysis showed that all viruses produced polyhedra of normal appearance.Green fluorescence can be apparently detected on the surface of the vBmBac(polh +)-EGFP polyhedra,but not the BmNPV polyhedra.Fluorescence analysis and anti-desiccation testing confirmed that EGFP was embedded in the polyhedra.As expected,the vBmBac(polh +)-LacZ polyhedra contained an amount of LacZ and had a higherβ-galactosidase activity.Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)and Western blotting were also performed to verify if the foreign proteins were immobilized into polyhedra.This study provides a new inspiration for efficient preservation of useful proteins and development of new pesticides with toxic proteins.
in the late phase of Bombyx mori nucleopolyhedrovirus （BmNPV） infection, a large amount of polyhedra appear in the infected cell nucleolus, these polyhedra being dense protein crystals protecting the incorporated virions from the harsh environment. To investigate whether the foreign protein could be immobilized into the polyhedra of BmNPV, two recombinant baculoviruses were generated by a novel BmNPV polyhedrin-plus （polh＋） Bac-to- Bac system, designated as vBmBac（polh＋）-enhanced green fluorescent protein （EGFP） and vBmBac（polh＋）-LacZ, which can express the polyhedrin and foreign protein simultaneously. Light microscopy analysis showed that all viruses produced polyhedra of normal appearance. Green fluorescence can be apparently detected on the surface of the vBmBac（polh＋）-EGFP polyhedra, but not the BmNPV polyhedra. Fluorescence analysis and anti-desiccation testing confirmed that EGFP was embedded in the polyhedra. As expected, the vBmBac（polh＋）-LacZ polyhedra contained an amount of LacZ and had a higher β-galactosidase activity. Sodium dodecyl sulfate polyacrylamide gel electrophoresis （SDS-PAGE） and Western blotting were also performed to verify if the foreign proteins were immobilized into polyhedra. This study provides a new inspiration for efficient preservation of useful proteins and development of new pesticides with toxic proteins.
基金
supported by the National Basic Research Program(973)of China(No.2012CB114600)
the Zhejiang Provincial Natural Science Foundation(No.Y3110058)
the Public Agricultural Program of Zhejiang Province(No.2012C32G2010076),China
