CORE--GENE TRANSFER

核心——基因转移

• 批准号: 6653346
• 负责人: Nader G. Abraham
• 金额: $ 31.48万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6653346
• 关键词: biomedical facility; biotechnology; recombinant virus; transfection; transfection /expression vector

Gene transfer technology either with the aid of viral vectors or in the form of naked DNA has been used extensively in recent years to elucidate mechanisms regulating vascular function and blood pressure. Preliminary data generated by project investigators with the help of the Core director indicate that this technology provides useful tools to each of the program project components. The core director collaborated with project investigators for more than ten years and recently produced and delivered recombinant retroviruses expression vectors and viruses (adenoviruses and retroviruses) containing primarily the CYP4A and HO genes for in vitro and in vivo studies proposed by the project investigators. The core will perform the following functions; 1) construct recombinant CYP4A, HO and COX-2 adenoviruses and retroviruses using commercially available vectors with and without GFP or beta-gal tagging; 2) adenoviruses and retroviruses using commercially available vectors with and without GRP or beta-gal tagging; 2) isolate and purify and propagate recombinant viral vectors; 3) determine titers and maintains stocks of recombinant viral vectors; 4) maintain stocks of the various plasmids and competent and packaging cells; 5) develop vectors carrying tissue and cell lineage specific promoters; and 6) provide the resources and expertise to each investigator for evaluating transgene expression and work along with the project investigators in validating transgene expression in vitro and in vivo using RT/PCR, Southern blot analysis and detection of GFP or beta-gal. Project investigators will be supported by the core director and the technical staff in the generation and production of molecular reagents, experimental design and data interpretation.

近年来，借助病毒载体或裸DNA形式的基因转移技术已被广泛用于阐明血管功能和血压的调节机制。项目调查人员在Core主管的帮助下生成的初步数据表明，该技术为每个项目组件提供了有用的工具。核心主任与项目研究者合作了十多年，最近生产并交付了项目研究者提出的主要含有CYP4A和HO基因的重组逆转录病毒表达载体和病毒（腺病毒和逆转录病毒），用于体外和体内研究。核心将执行以下功能；1)利用市售载体构建重组CYP4A、HO和COX-2腺病毒和逆转录病毒，有或没有GFP或β -gal标记；2)腺病毒和逆转录病毒使用市售载体，有或没有GRP或β -gal标记；2)分离纯化并繁殖重组病毒载体；3)确定病毒滴度并维持重组病毒载体的库存；4)保持各种质粒、活性细胞和包装细胞的库存；5)开发携带组织和细胞谱系特异性启动子的载体；6)为每位研究者提供评估转基因表达的资源和专业知识，并与项目研究者一起使用RT/PCR、Southern blot分析和GFP或β -gal检测来验证转基因在体外和体内的表达。项目研究者将在核心主任和技术人员的支持下进行分子试剂的生成和制作、实验设计和数据解释。