中文摘要：

目的构建人铜锌超氧化物歧化酶基因（Cu/Zn-SOD,SOD1）启动子驱动的红色荧光蛋白报告基因载体,并对其功能进行鉴定,为研究细胞内SOD1基因表达调控和相关的信号转导机制提供重要工具。方法提取人基因组DNA,PCR扩增SOD1 5′非编码区启动子序列（-1 499-＋17）;采用基因重组技术构建由SOD1启动子驱动的红色荧光蛋白报告基因载体,将该载体瞬时转染Hella细胞,观察静息或佛波酯（PMA）刺激下红色荧光蛋白表达,以检测SOD1启动子的转录活性及其基因表达。结果酶切和DNA测序证明所构建红色荧光蛋白报告基因载体正确;该表达载体静息状态下在Hella细胞表达少量红色荧光蛋白,经PMA刺激后,能够表达较多高亮度的红色荧光蛋白。结论成功构建了SOD1启动子驱动的红色荧光蛋白报告基因载体,其对相关刺激有很好的反应,可用于SOD1基因表达信号调控机制的研究。

英文摘要：

Objective To construct the red fluorescent protein reporter gene plasmid driven by human Cu/Zn superoxide dismutase gene （Cu/Zn-SOD, SOD1） promoter, to study the function of the reporter gene, and to provide a tool for the further study on the regulation mechanism of the expression of SOD1 and related signal transduction pathways. Methods Genomic DNA was extracted from human blood cells. The promoter sequence （-1499-＋17） in 5′ non-coding region of SOD1 was amplified by polymerase chain reaction （PCR） method. The red fluorescent protein reporter gene vector driven by human SOD1 promoter was construeted by gene recombination technique. The recombined vector was transiently transfected into Hella cells, and the expression of the red fluorescent protein in Hella cells was then observed by fluorescent microscopy when the calls were at rest or stimulated by Phorbol-12 myristate-13-acetate （PMA）, by which the transcription activity of the SOD1 promoter and the expression of SOD1 were determined. Results The red fluorescent protein reporter gene vector was correctly constructed, which was proved by restriction enzyme digestion and DNA sequencing. The vector expressed a little of red fluorescent protein in Hella cells at rest, but more red fluorescent protein was strongly expressed in the cells after PMA stimulation. Conclusions A red fluorescent protein reporter gene plasmid driven by human SOD1 promoter has been successfully constructed, and the plasmid responds well to related stimulus. The reporter gene vector will provide an efficient and convenient tool for the further study on the regulation mechanism of the expression of SOD1.