CORE--GENE TRANSFER

核心——基因转移

• 批准号: 6808277
• 负责人: PAUL J MCDERMOTT
• 金额: $ 6.5万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6808277
• 关键词: Adenoviridae; biomedical facility; genetic manipulation; heart enlargement; intracardiac pressure; microorganism resource /registry; recombinant virus; transfection

The function of the Gene Transfer Core is to provide a facility for the construction, screening, large-scale purification, and titrating of replication-defective, recombinant adenoviruses. Recombinant adenoviruses provide a reliable method for high efficiency gene transfer into adult cardiocytes and for directly altering the phenotype. Specifically, the core will 1) maintain lines of human 293 endothelial kidney cells [HEK], 2) amplify and maintain purified stocks of adenovirus shuttle plasmids and adenovirus backbone plasmids, 3) transfect recombinant adenoviral DNA into 293 HEK cells, 4) propagate recombinant adenoviruses in 293 HEK cells, 5) plaque-purify recombinant adenoviruses and perform large-scale purifications, and 6) titer purified adenoviruses by plaque assays in 293 HEK cells. The core will also be responsible for long-term storage of recombinant adenoviruses and will assist individual investigators in carrying out gene transfer protocols in adult cardiocytes. A full-time research specialist has been trained in all of the procedures and techniques involved in running this core, and will be supervised by the core director.

基因转移核心的功能是为复制缺陷型重组腺病毒的构建、筛选、大规模纯化和滴定提供设施。重组腺病毒提供了一种可靠的方法，高效基因转移到成人心肌细胞和直接改变表型。具体而言，核心将1）维持人293内皮肾细胞[HEK]的细胞系，2）扩增和维持腺病毒穿梭质粒和腺病毒骨架质粒的纯化原液，3）将重组腺病毒DNA转染到293 HEK细胞中，4）在293 HEK细胞中繁殖重组腺病毒，5）噬斑纯化重组腺病毒并进行大规模纯化，和6）在293 HEK细胞中通过噬斑测定来滴定纯化的腺病毒。该核心还将负责重组腺病毒的长期储存，并将协助个别研究人员在成人心肌细胞中进行基因转移方案。一名全职研究专家已接受了运行该核心所涉及的所有程序和技术的培训，并将接受核心主任的监督。