J亚群禽白血病病毒SUJ及兔IgGFc基因在腺病毒表达系统中的融合表达  

Fusion expression of subgroup J avian leukosis virus gp85 gene with rabbit Ig GFc gene in adenovirus expression system and its application

作　　者：梅梅[1,2,3] 钱科[4] 唐应华[1,3] 秦爱建[2,3] 侯继波[1,3] 

机构地区：[1]江苏省农业科学院,江苏南京210014 [2]扬州大学江苏省动物预防医学重点实验室,江苏扬州225009 [3]江苏省动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009 [4]江苏省出入境检验检疫局,江苏南京210001

出　　处：《江苏农业学报》2016年第4期848-853,共6页Jiangsu Journal of Agricultural Sciences

基　　金：国家自然科学基金青年基金项目(31201881);教育部创新团队项目(IRT0978);江苏省普通高校研究生科研创新计划项目(CXL2_0938X1)

摘　　要：为获得J亚群禽白血病病毒(ALV-J)囊膜蛋白gp85和兔IgGFc的融合蛋白,通过腺病毒表达系统融合表达ALV-J SUJ和兔IgGFc(r IgGFc)基因。将pc DNA3.1-SUJ-r IgGFc用XhoⅠ、KpnⅠ进行双酶切,获得SUJ-r IgGFc基因,将其克隆至p Shuttle-CMV质粒,构建穿梭载体p Shuttle-CMV-SUJ-r IgGFc,重组质粒转化含p Adeasy-1的BJ5183-AD-1感受态细胞,获得重组腺病毒质粒p Ad-SUJ-r IgGFc,最后转染人293T细胞。表达的融合蛋白可被ALV-J单克隆抗体JE9以及羊抗兔IgG识别,重组蛋白的分子量大小约9.5×104,且与JE9及羊抗兔IgG都有很好的反应性;免疫共沉淀试验结果显示融合蛋白可与DF1细胞膜蛋白混合物反应并获得差异蛋白。该融合蛋白可与ALV-J宿主细胞蛋白发生特异性反应。To obtain the fusion protein of subgroup J avian leukosis virus( ALV-J) envelope protein gp85 and rabbit IgGFc,ALV-J gp85 gene fused with rabbit IgGFc was expressed by adenovirus expression system. The ALV-J gp85( SU)and rabbit IgGFc( r IgGFc) gene were digested from plasmid pc DNA3. 1-SUJ-r IgGFc with XhoⅠand KpnⅠand cloned into p Shuttle-CMV to construct the recombinant plasmid p Shuttle-CMV-SUJ-r IgGFc which was transformed into BJ5183-AD-1competent cells containing p Adeasy-1 to construct the recombinant adenovirus plasmid p Ad-SUJ-r IgGFc. The recombinant adenovirus was obtained by transfecting the recombinant plasmid p Ad-SUJ-r IgGFc into 293 T cells. The fusion protein can be recognized by m Ab JE9 specific to gp85 of ALV-J and antibodies against rabbit IgG with the molecular weight about9. 5 × 104. The result of Co-IP showed that the fusion protein SUJ-r IgGFc reacted with the membrane proteins from DF1 cells and produced differential proteins,indicative of the specific reaction between fused protein and the protein from host cells of ALV-J.

关 键 词：J亚群禽白血病病毒 GP85基因 兔IgGFc 腺病毒 融合表达 

分 类 号：S852.65[农业科学—基础兽医学]