重组腺病毒载体含人抑瘤素的扩增及纯化

The Propagation and Purification of Human HOSM Recombinant Adenovirus Vector

目的 :构建含人抑瘤素 (HOSM)基因的重组腺病毒载体 ,观察其对肿瘤细胞生长的影响。方法 :用脂质体介导质粒DNA同源重组方法构建HOSM基因的重组缺陷型腺病毒载体 ,用聚合酶链反应 (PCR)法鉴定所构建的ad HOSM载体 ,在人胚肾细胞 (2 93细胞 )中扩增病毒。CsCl梯度超速离心纯化病毒 ,噬斑法测定病毒的滴度。结果 :获得了高纯度、高滴度的重组腺病毒载体。结论 :细胞内质粒DNA同源重组法能有效构建重组腺病毒载体 ,2 93细胞能有效扩增病毒。CsCl梯度超速离心法能制备高纯度的病毒 。

Objective: To construct human HOSM recombinant adenoviral vector and study its effect on the growth of tumor cells. Methods: Plasmid DNA homologous recombination modality was used to construct the replication deficient adenoviral vector encoding human HOSM gene. The ad hosm were propagated in cell line 293, purified by CsCl gradient ultracentrifugation and titered by plaque assay. Results: The recombinant adenoviral vectors were harvested with high titer and purity. Conclusions: The recombinant adenoviral vector can be constructed efficiently with homologous recombination in vivo and adenovirus can be propagated effectively in cell line 293. Adenovirus with high purity can be harvested by CsCl gradient ultracentrifugation. This research provides a foundation to the future work applying the recombinant adenoviral vector encoding hosm gene in studies of biological activities of tumor cells.