不同启动子在昆虫杆状病毒表达系统中的活性分析.docVIP

生物工程学报 Chin J Biotech 2008, April 25; 24(4: 598-603 Chinese Journal of Biotechnology ISSN 1000-3061 cjb@ ? 2008 Institute of Microbiology, CAS CSM, All rights reserved 不同启动子在昆虫杆状病毒表达系统中的活性分析 王煜1, 2, 高辉1, 陈川1, 赵和平1, 李淼1, 孙元1, 仇华吉1 1 中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室猪传染病研究室, 哈尔滨 150001 2 内蒙古农业大学动物科学与医学学院, 呼和浩特 010018 摘 要: 为了比较不同启动子在杆状病毒/昆虫细胞中的活性, 利用对虾白斑综合症病毒(White spot syndrome virus, WSSV 的ie1启动子及其截短的mie1启动子、杆状病毒ETL 启动子及其加长的mETL 启动子以及杆状病毒多角体启动子P PH , 构建了含有不同启动子控制下的EGFP 报告基因的重组杆状病毒, 分别感染Sf9昆虫细胞, 利用流式细胞术检测报告基因的表达水平。结果表明, WSSV的ie1启动子和杆状病毒的mETL 启动子在昆虫细胞Sf9细胞中都具有较强而早的启动子活性, 能够控制报告基因早期高效表达, 而P PH 在感染后期才表现较强活性。并且研究中发现, 杆状病毒同源重复区(hr1可以增强ETL 启动子的活性。 关键词: 重组杆状病毒, 启动子活性, 流式细胞术 Promoter Activity of Different Promoters in Recombinant Baculovirus-infected Sf9 Cells Yu Wang1, 2, Hui Gao1, Chuan Chen1, Heping Zhao1, Miao Li1, Yuan Sun1, and Huaji Qiu1 1 Division of Swine Infectious Diseases, National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China 2 College of Animal Sciences and Veterinary Medicine, Inner Mongolian Agricultural University, Hohhot 010018, China Abstract : To compare the activity of different promoter in baculovirus-insect system, a series of recombinant baculoviruses were generated harboring the E-GFP reporter gene under the control of one of 5 promoters, including the ie1 promoter of shrimp white spot syndrome virus (WSSV, the truncated ie1 (mie1 promoter, the ETL promoter of the baculovirus, the elongated ETL (mETL promoter, and the polyhedron promoter (PPH of the baculovirus. The expression efficiency of the E-GFP reporter gene in the recombinant baculovirus-infected Sf9 cells was determined by flow cytometry. The results showed that both ie1 and mETL promoters had a strong promoter activity at early phase, while PPH showed a strong promoter activity at late phase. The ie1 promoter suggested the strongest promoter activity. The homologous region 1 (hr1 was also found t