目的 构建S100P基因短发夹RNA(shRNA)慢病毒载体 ,并在胃癌细胞中鉴定其沉默效率.方法 设计S100P基因特异性的RNA干扰序列 ,构建shRNA慢病毒载体 ,与S100P过表达质粒共转染293T细胞.随后筛选有效的shRNA慢病毒载体 ,与pHelper 1 .0和pHelper 2 .0质粒共转染293T细胞 ,包装病毒 ,感染胃癌细胞株MGC-803和SGC-7901.采用RT-PCR和Western blot检测S100P基因的敲减效率.结果 成功构建S100P基因的shRNA慢病毒载体 ,MGC-803和SGC-7901细胞中S100P基因的mRNA和蛋白表达均降低(P<0 .05).结论 成功构建的S100P基因shRNA慢病毒载体能够在细胞水平有效沉默靶基因.%Objective To construct a short hairpin RNA (shRNA ) lentiviral vector targeting human S100P gene and identify its silence effect in gastric cancer cells .Methods Five specific siRNA sequences targeting human S100P gene were designed and cloned into GV115-GFP lentiviral vectors , which were co-transfected with overexpressing vector of S100P into 293T cells .The valid shRNA lentiviral vectors were screened and transfected with pHelper 1 .0 and pHelper 2 .0 plasmids into 293T cells for virus packaging .Then gastric cancer cell lines MGC-803 and SGC-7901 were infected with specific shRNA lentivirus ,and the mRNA and protein expressions of S100P gene were detected by RT-PCR and Western blot ,respectively .Results shRNA lentiviral vector targeting human S 100P gene was successfully constructed ,and the mRNA and protein expressions of S100P gene were significantly decreased in MGC-803 and SGC-7901 cells (P<0 .05) .Conclusion The recombinant shRNA lentiviral vector targeting S 100P gene can effectively silence target gene at the cellular level .
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