马立克氏病病毒pp38基因和1.8kb转录子之间双向启动子的特性研究  

Study on the characterization of the bi-directional promoter betweenpp38 gene and 1.8kb mRNA transcripts ofMarek's disease viruses

作　　者：丁家波[1] 崔治中[1] 姜世金[1] 孙爱军[1] 孙淑红[1] 

机构地区：[1]山东农业大学动物科技学院,泰安271018

出　　处：《微生物学报》2005年第3期363-367,共5页Acta Microbiologica Sinica

基　　金：国家自然科学基金资助项目 ( 3 0 0 70 5 44 )~~

摘　　要：从马立克氏病病毒(MDV)基因组DNA复制原点区某一点,将介于MDVpp38基因和1 8kb转录子之间的双向启动子分割成两个单方向的启动子。以pp38为报告基因,pUC18质粒为载体,构建了含不同方向完整启动子序列的pProfpp38和pProrpp38质粒,以及含分割后单方向启动子序列的pdProfpp38和pdProrpp38质粒。4种质粒分别转染鸡胚成纤维细胞(Chickenembryofibroblast,CEF)后,均能检测到pp38基因的表达。进一步以氯霉素乙酰转移酶(Chloramphenicolacetyltransferase,CAT)为报告基因,构建了含不同方向完整双向启动子的pProfCAT和pProrCAT质粒,以及含分割后单方向启动子序列的pdProfCAT和pdProrCAT质粒。通过转染试验,定量分析了完整启动子和分割后启动子在两个方向上的启动活性。实验结果表明,分割后的启动子在两个方向上的启动活性均比相应方向上完整启动子的活性低,其中1 8kb转录子方向上的活性下降了4The bi-directional promoter betweenpp38 gene and 1.8kb mRNA transcripts ofMareks disease viruses (MDV) was divided into two single_direction promoters from the replication of MDV genomic DNA. The pp38 gene was cloned into pUC18 vector for plasmid pUC_pp38. Then the complete bi_directional promoter was cloned into pUC_pp38 in two directions to form plasmids pPro f pp38 and pPro r pp38, and the divided two single directional promoters were cloned in pUC_pp38 for plasmids pdPro f pp38 and pdPro r pp38. 24 to 48 hours after transfection to chicken embryo fibroblast (CEF) cells, the expression ofpp38 could be detected in above 4 samples with Indirect Immuno_fluorescent Assay (IFA). In order to analysis the activity of the promoter quantificationally, CAT was used as the report gene. The complete or divided promoters were cloned into pCAT_Basic vector for plasmids pPro f CAT, pPro r CAT, pdPro f CAT and pdPro r CAT. The activity of CAT was measured from the lysed CEF cells, when they were transfected for 48 hours by the above four plasmids, respectively. The results showed the activity of the divided promoters reduce on both directions, especially for the direction of 1.8kb mRNA transcript, nearly down to 1/41.

关 键 词：马立克氏病病毒 PP38基因 1.8kb转录子 双向启动子 

分 类 号：S852.65[农业科学—基础兽医学]