乙型肝炎病毒e抗原结合蛋白基因的筛选  

Screening of the genes of hepatitis B virus e antigen interacting proteins

作　　者：李伯安[1] 舒翠莉[1] 侯俊[1] 戚杨[1] 李靖[1] 何卫平[1] 程云[1] 

机构地区：[1]解放军第三○二医院免疫室,北京100039

出　　处：《中华实验和临床病毒学杂志》2005年第3期271-274,共4页Chinese Journal of Experimental and Clinical Virology

基　　金：国家自然科学基金(30100170)

摘　　要：目的筛选并克隆人肝细胞中与乙型肝炎病毒e抗原(HBeAg)相互作用的蛋白质基因。方法将重组诱饵质粒pGBKT7eAg转化酵母细胞AH109后与预转了人肝cDNA文库质粒的酵母细胞Y187进行配合,在营养缺陷型培养基上进行双重筛选阳性菌落,提取质粒后转化大肠埃希菌并进行序列测定,进行生物信息学分析。结果配合后筛选出既能在4缺(SD/TrpLeuAdeHis)培养基又能在铺有Xα半乳糖(Xαgal)的4缺培养基上生长并变成蓝色的真阳性菌落245株。完成了101株克隆的测定,经过序列分析排除了重复同源序列后,最终确定其中有41株不同的基因,其中人类同源基因35条,其余6株为未知基因。结论成功克隆出肝细胞中的HBeAg结合蛋白基因,为进一步研究HBeAg的功能提供了新线索。Objective To screen and clone the genes in hepatocytes which encode protein that can interact with hepatitis B e antigen（HBeAg） by yeast-two hybridization. Methods Recombined HBeAg bait plasmid（pGBKT7-eAg） was transformed into yeast AH 109, followed by mating with yeast Y187 containing liver cDNA library plasmid in 2 × YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium （SD/-Trp-Leu-Ade-His） which contains X- α-gal for selecting positive blue clones. Then positive clones were selected and plasmids were prepared and sequenced. Finally, bioinforamatics analysis was performed. Results Tolally 245 positive colonies were selected and 101 colonies were sequenced. Through sequences alignment, 6 novel genes and 35 recorded genes were screened. Conclusion Genes of HBeAg interacting proteins have been cloned successfully, which brings some new clues for further studies on the biological functions of HBeAg and the related proteins.

关 键 词：肝炎e抗原 乙型 基因 酵母菌 杂交 乙型肝炎病毒E抗原 结合蛋白基因 筛选 生物信息学分析 HBEAG 人肝细胞 

分 类 号：R392[医药卫生—免疫学]