项目名称： 单颗粒病毒实时示踪方法研究杆状病毒AcMNPV转导不同组织来源人源细胞的机制

项目编号： No.31500131

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 生物科学

项目作者： 郑方亮

作者单位： 辽宁大学

项目金额： 23万元

中文摘要： 目前，虽然基因治疗已经用于某些疾病的临床治疗，但是该技术的安全性和有效性仍有待进一步提升。基因治疗的核心技术之一是获得安全、高效的基因转移载体。杆状病毒是以节肢动物为唯一宿主的昆虫病毒，它可以侵入哺乳动物细胞，表达所携带的外源基因，但却不能在其中复制，这为将其用于基因转移的载体提供了可靠的安全保障。然而，杆状病毒转导哺乳动物细胞的具体机制尚不明确，这很大程度上限制了其在基因治疗上的应用。为了解析这一机制，项目组通过基因工程改造和化学标记等方法，成功构建了SA-QDs和/或CFP标记的重组杆状病毒AcMNPV，并拟利用此重组病毒转导不同来源的人源细胞，通过实时观察荧光的分离和运动轨迹来探索转导后病毒与宿主膜结构的融合途径、病毒粒子在细胞内的运动轨迹以及病毒核酸的释放位点，阐明杆状病毒转导哺乳动物细胞的机制，为推动并完善杆状病毒转基因载体和工程细胞的设计以及尽快实现临床应用提供重要的理论依据。

中文关键词： 杆状病毒；量子点；示踪

英文摘要： At present, Although gene therapy is used for clinical treatment of some diseases, the safety and effectivity of this technique remains to be promoted. One of the core technology of gene therapy is to acquire an safe and effective gene transfer vector. Baculoviruses are highly restricted insect viruses, which can enter, express foreign gene, but not replicate in mammalian cells, that supply the virus insurance for being used as gene transfer vector. However, mechanisms of baculovirus transduction are poorly understood, that limit its application in gene therapy to a large extent. To analyse the mechanism, we constructed SA-QDs and/or CFP labeled recombinant baculovirus AcMNPV by genomic engineering and chemical labeling, and then, by real-time tracking SA-QDs and/or CFP-labelled virus in cells originated from different human organs, we will focus on the following research questions: how and where does membrane fusion take place during virus entry (membrane fusion mechanism)? Which subcellular skeleton protien(s) and organ(s) participate in virus transportation within the cells (virus transportation machanism)? Where does virus release its DNA (DNA release site)? By elusidating the mechanisms above, this study will provide theoretical basis for development of gene transfer vectors, gene engineering cells and the applications of baculovirus in gene therapy.

英文关键词： Baculovirus;QDs;Tracing