禽马立克氏病毒糖蛋白B基因在家蚕中的表达

将马克克氏病毒（Marek'sdiseasevirus，MDV）糖蛋白B（gB）基因克隆入转移载体pBac－PAK8中，得到重组转移载体质粒pBacPAK（gB）。经限制性酶切图谱结合Southernblot分析鉴定表明，gB基因以正确方向插入转移载体，受多角体蛋白基因启动子控制。将此转移载体与经CvnI酶切线性化的亲本病毒Bm－BacPAK6DNA通过脂质体法共转梁家蚕细胞，只有发生重组的病毒才有复制增殖的能力。然后通过蓝白斑筛选、结合点杂交，纯化得到重组的空斑病毒vBM，用该重组病毒接种家蚕5龄幼虫，对表达产物进行SDS－PAGE分析，检测到gB基因在家蚕中高效表达，表达产物的分子量主要为97KD，表达量约为1mg／mL血淋巴。

EXPRESSING MAREK'S DISEASE VIRUS GLYCOPROTEIN B GENE IN SILKWORM

In this study, Marek's disease virus (MDV) Glycoprotein B(gB) gene was clonedinto transfer vector pBacPAK8 and the recombinant transfer vector pBacPAK (gB) wasobtained. The analysis with restriction endonuclease digestion and Southern blot showedthat gB gene was inserted into the transfer vector in the right direction and controlled bythe polyhedrin promoter. This recombinant transfer vector and parental virus Bm-Bac-PAK6 DNA (Cleaved with Cvn I ) were used to cotransfect Bm-N Cells with lipofectin.Only the recombinant viruses could replicate in cells. After that, the recombinant virusplaque was selected and purified by blue-white plaque and dot hybridization assay. Thefifth instar larvae of Bombyx mori were injected with the purified recombinant virus.SDS-PAGE analysis of products from the recombinant virus infected silkworms showedthat MDV gB gene was expressed at high efficiency. The molecular weight of the ex-pressed products is mainly 97kD, the expressed level is 1 mg per ml hemolymph.