CORE--VECTOR 2--RETROVIRUS

核心--载体2--逆转录病毒

• 批准号: 6501119
• 负责人: MICHEL SADELAIN
• 金额: $ 12.63万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-01 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6501119
• 关键词: Retroviridae; biomedical facility; cell line; microorganism culture; southern blotting; transfection; transfection /expression vector

Recombinant retroviruses are powerful tools for gene transfer that are used in experimental as well as clinical applications. Retroviral-mediated gene transfer leads to stable proviral integration in the target-cell genome, a key feature when foreign genetic material must be reliably transmitted to the progeny of transduced progenitor cells or clonogenic cells. The projects that make up this Program Project rely in part or exclusively on retroviral-mediated gene transfer to modify primary cells prepared from heart, lung, blood, bone marrow, and embryonic stem cells. The construction of a retroviral vector and the generation of a stable high-titer producer cell line are elaborate procedures with well- defined Checkpoints that are best handled in a highly specialized core laboratory. Centralized generation of high-quality producer cell lines and retroviral stocks will be cost effective, permit standardization of procedures, and facilitate the exchange of information and expertise between the different projects. The specific aims of the Retrovirus Core are: 1. construction of the recombinant retroviral vectors; 2. transfection of vector DNA in packaging cells and selection of producer cell lines; 3. characterization of vector transmission in standard cell lines by Southern blot analysis; 4. expansion of independent clones and identification of the best clone for specific target cells;5. titration of cell-free retroviral stocks; 6. detection of replication-competent retrovirus in cell-free viral stocks. The Retrovirus Core is directed by M. Sadelain, M.D., Ph.D., who is assisted by a senior research technician with considerable experience in cell culture and molecular techniques.

重组逆转录病毒是用于基因转移的强大工具 用于实验和临床应用中。 逆转录病毒介导的基因转移导致病毒稳定 集成在目标细胞基因组中，这是外国时的关键特征 遗传物质必须可靠地传播到 转导的祖细胞或克隆性细胞。这些项目 组成这个程序项目部分依靠或仅仅依靠 逆转录病毒介导的基因转移以修饰制备的原代细胞 从心脏，肺，血液，骨髓和胚胎干细胞中。这 逆转录病毒载体的构建和稳定的产生 高敏机生产者细胞系是精心制作的程序 定义的检查点最好在高度专业化中处理 核心实验室。集中的高质量生产商 细胞系和逆转录病毒库存将具有成本效益，许可证 程序的标准化，并促进交换 不同项目之间的信息和专业知识。这 逆转录病毒核心的具体目的是：1。 重组逆转录病毒载体； 2。向量DNA中的转染 包装细胞和生产者细胞系的选择； 3。 标准单元线中向量传输的表征 Southern印迹分析； 4。独立克隆的扩展和 鉴定特定靶细胞的最佳克隆; 5。 无细胞逆转录病毒量的滴定； 6。检测 无细胞病毒量的复制逆转录病毒。这 逆转录病毒核心由M. Sadelain博士执导 在高级研究技术员的协助下 在细胞培养和分子技术方面的经验。