中文摘要：

构建可表达增强型绿色荧光蛋白（Enhanced green fluorescent protein，EGFP）的辅助病毒依赖型腺病毒载体（Helper-dependent adenoviral vector，HDAd），并完成大量制备、纯化和体外表达鉴定。荧光显微镜证实HDAd／EGFP可表达，电镜下观察到经CsCl纯化后的腺病毒的典型形态。分光光度计法测定病毒的浓度为4．0×10坦颗粒数（Virus particle，vp）／mL。与可表达EGFP的第一代腺病毒载体（First generation adenoviral vector，FGAd）FGAd／EGFP进行了体外感染和转基因表达效率的比较研究，分别用约2000vp／细胞的HDAd／EGFP和FGAd／EGFP感染A549细胞，流式细胞仪检测EGFP的表达情况。通过相同时间点流式细胞仪分析EGFP的表达情况，可见HDAd／EGFP感染早期的A549细胞较FGAd／EGFP有更高的荧光表达率及更高的表达强度，显示HDAd载体具有转基因瞬时高表达的特性，是一种更有价值的疫苗载体。

英文摘要：

To investigate the transgenic expressing efficacy of helper-dependent adenoviral vector （HDAd） in vitro, we constructed a HDAd encoding enhanced green fluorescent protein （EGFP）, denominated as HDAd/EGFP, performed large scale preparation and purification, and then identified the purified HDAd/EGFP under fluorescent microscope and electron microscope. After the concentration of HDAd/EGFP was determined by spectrophotometer, the transgenic expression efficiency of HDAd/EGFP was compared with first generation adenoviral vector encoding EGFP （FGAd/EGFP） in vitro. Therefore, we infected A549 cells with 2 000 virus particles （vp） per cell by HDAd/EGFP and FGAd/EGFP respectively and analyzed EGFP expressing level by flow cytometry. Consequently, the fluorescent expression rate and fluorescent intensity of EGFP were higher in early infected A549 cells by HDAd/EGFP than by FGAd/EGFP. HDAd, capable of expressing transgene instantly and efficiently in vitro, is a potential vaccine vector.