HCMV IE86对ARPE-19细胞周期的影响  

Effects of HCMV IE86 on Cell Cycles in ARPE-19 Cells

作　　者：刘海燕[1] 李玲[1] 白志强[1] 王斌[1] 钱冬萌[1] 丁守怡[1] 宋旭霞[1] 赵巍[1] 闫志勇[1] 苏洁[1] 杨丽[1] 

机构地区：[1]青岛大学医学院微生物教研室,山东青岛266071

出　　处：《现代生物医学进展》2009年第17期3219-3222,共4页Progress in Modern Biomedicine

基　　金：国家自然科学基金资助项目(30770105);青岛市科技计划项目(08-1-3-30-jch);泰山学者建设工程专项经费资助

摘　　要：目的:构建真核表达质粒pEGFP-N3-IE2,瞬时转染ARPE-19细胞,探讨其对视网膜色素上皮细胞周期的影响。方法:采用PCR方法从质粒pIEP86AD169扩增出IE2片段,将其定向克隆于pEGFP-N3,构建真核表达质粒pEGFP-N3-IE2,酶切、测序鉴定,脂质体介导瞬时转染ARPE-19细胞,RT-PCR、免疫荧光法分析IE2基因在mRNA和蛋白水平的表达,流式细胞仪检测转染前后细胞周期的变化。结果:成功构建重组质粒pEGFP-N3-IE2,并在被转染细胞中检测到IE2基因的表达;瞬时转染后细胞周期表现为S期细胞比例显著增高。结论:成功构建pEGFP-N3-IE2真核表达质粒,重组质粒能在ARPE-19细胞中顺利表达IE2,IE2基因可以引起ARPE-19细胞的细胞周期S期阻滞。Objective： Constructing eukaryotic expression vector of HCMV IE2 and express in ARPE-19 cells after transient trans-fection in order to investigate the effect on cell cycles. Methods： The HCMV IE2 gene was amplified from plasmid plEP86AD169 by PCR, and cloned to eukaryotic expression vector pEGFP-N3. The recombinant plasmid pEGFP-N3-IE2 was transfected into ARPE-19 cells with Lipofectamine 2000. The expression of IE2 was tested in ARPE-19 cells by RT-PCR and immunofluorescence assay. The cell cycle was analyzed by FCM. Results： The recombinant plasmid pEGFP-N3-IE2 had been constructed correctly and the expression of IE2 gene could be detected in APRE-19 cells. The FCM analytic results showed that HCMV IE286 could increase the S-phase cell number in APRE-19 cells. Conclusion： The HCMV IE286 is successfully subcloned into eukaryotic expression vector and expressed in ARPE-19 cells. It can induce ARPE-19 cell cycle S arrest.

关 键 词：IE2 HCMV ARPE-19 细胞周期 

分 类 号：R774.11[医药卫生—眼科]