中文摘要：

目的 构建以SM22α启动子驱动EGFP表达的慢病毒载体,研究此载体在血管平滑肌细胞（vascular smooth muscle cells,VSMCs）中表达的效率和特异性.方法 从小鼠的基因组中采用PCR扩增SM22α启动子,将目的基因连接至T载体,并进行测序鉴定,将目的基因重组至pGC-FU载体,三质粒系统采用脂质体介导转染293T细胞进行病毒包装,体外感染VSMCs,大鼠动脉内皮细胞,人乳腺癌细胞系SK-BR-3,荧光显微镜观察EGFP的表达情况.结果 PCR扩增得到的SM22α启动子经测序证实正确;成功构建了SM22α启动子驱动EGFP的慢病毒载体LV-SM22α-EGFP,SM22α启动子能调控EGFP在VSMCs中高效表达,感染效率在95%以上,在血管内皮细胞和人乳腺癌细胞系SK-BR-3中不表达,对照慢病毒组EGFP在三种细胞中均表达,在VSMCs的感染效率在30%左右,且表达较低.结论 SM22α启动子能够特异性驱动EGFP在VSMCs中表达,LV-SM22α-EGFP能作为研究血管疾病的良好基因转移载体.

英文摘要：

Objective To construct lentiviral vector carrying EGFP driven by SM22α pro- moter and to explore the specificity and efficiency of EGFP gene expression in VSMCs infected by this ]entiviral vector. Methods SM22α promoter was amplified from mouse genome by PCR technique, the target gene was ligated to T vector, and identified by sequencing. The target gene was reconstructed to pGC - FU vector, Lipofectamine 2000 - mediated tri - vector system was transfected into 293T cells, and lentivial vector was packaged to infect human breast cancer cell line SK - BR - 3, VSMCs and endothelial cells from rat aorta. The expression of EGFP was observed under the fluorescence microscopy. Results The SM22α promoter gene amplified by PCR was identified to be correct by sequencing. The lentivirus driven by SM22α promoter was constructed successfully, the SM22α promoter could drive high efficacy expression of EGFP exclusive in VSMCs, and the transfection efficacy was more than 95%. The report gene EGFP could be expressed in three kinds of cells by infection of control common lentivirus, and the transfection efficacy in VSMCs was about 30% and the expression level of EGFP was lower. Conclusion The SM22α promoter can drive EGFP specifically expression in vascular smooth muscle cells, LV - SM22 - EGFP may serve as a optimal gene delivery vector for vascular disease research.