中文摘要：

目的制备环氧化酶-2（COX-2）的RNA干扰（RNAi）重组慢病毒（lentivirus）,观察COX-2表达沉默对人食管鳞癌（ESCC）细胞增殖的影响。方法将目的基因短发卡RNA（sh RNA）载体与包装载体共转染293T细胞,收集上清液后浓缩纯化、滴度测定、感染ESCC细胞;应用荧光显微镜观察包装效率及感染效率,实时荧光定量PCR及Western blot评价COX-2基因沉默效应,MTS法检测细胞的增殖。结果成功制备了沉默COX-2的重组慢病毒,该病毒感染ESCC细胞后,COX-2 m RNA和蛋白的表达量分别降低了80%和50%左右,细胞的增殖明显减慢。结论慢病毒介导的sh RNA干扰能有效沉默ESCC细胞COX-2基因,COX-2表达沉默抑制ESCC细胞增殖。

英文摘要：

ObjectiveTo construct a recombinant lentivirus for RNA interference （RNAi） of COX-2 gene and observe its effect on esophagealsquamous-cell carcinoma （ESCC） cell proliferation. MethodsThe COX-2 short hairpin RNA （shRNA） and packaging vectors were cotransfected to 293T cells to produce the recombinant lentivirus, which were further ultra-centrifuged, tittered and used to infect ESCC cells. The transfection efficiency and infection efficiency were evaluated with fluorescence microscope. Real-time PCR and Western blot were used to confirm the silencing of COX-2. MTS assay was used to study the effect of COX-2 on cell proliferation.ResultsWe successfully prepared the recombinant lentivirus for RNAi of COX-2 gene. The expression of COX-2 mRNA and protein were effectively silenced and the proliferation of COX-2 silenced cells was significantly inhibited.ConclusionLentivirus mediated shRNA effectively inhibited the expression of COX-2 and the proliferation ability of human ESCC cells.