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目的 :阐明凋亡抑制蛋白存活素 (survivin)的基因表达调控机制。方法 :采用PCR的方法从人HeLaS3细胞基因组DNA中扩增得到survivin基因上、下游侧翼序列 ,将它们分别克隆至报告基因表达载体中 ,依据报告基因的瞬时表达结果确定它们对survivin基因的表达调控作用 ,进而采用定向缺失的方法确定其核心启动子 ,并通过凝胶阻滞实验进行验证。结果与结论 :survivin基因起始密码子上游 2 6 8bp的序列是指导其在肿瘤细胞中周期性表达的最小核心启动子 (mini_promoter)。此外 ,体外培养的正常人外周血淋巴细胞经PHA_P与IL_2的刺激增殖的同时 ,survivin基因也得以表达 ,提示survivin基因的表达与细胞的增殖过程密切相关。
Objective: To elucidate the regulatory mechanism of the gene expression of survivin. Methods: The upstream and downstream flanking sequences of survivin gene were amplified by PCR from human HeLaS3 cells and cloned into the expression vector of reporter gene. The expression of survivin gene was determined according to the transient expression of reporter gene And then use directional deletion method to determine its core promoter, and verified by gel retardation experiment. RESULTS AND CONCLUSION: The 268 bp sequence upstream of the start codon of survivin gene is the smallest promoter for cyclical expression in tumor cells (mini_promoter). In addition, normal human peripheral blood lymphocytes cultured in vitro stimulated by PHA_P and IL_2 proliferation, survivin gene expression, suggesting that the expression of survivin gene and cell proliferation is closely related.