Szulc Jolanta, Aebischer Patrick
Neurosciences Institute, Swiss Federal Institute of Technology Lausanne, Lausanne, Switzerland.
Methods Mol Biol. 2008;434:291-309. doi: 10.1007/978-1-60327-248-3_18.
Drug-inducible systems allowing the control of transgene expression and knockdown in mammalian cells are invaluable tools for genetic research, and could also play important roles in translational research or gene therapy. We and others have developed a lentivector-based, conditional gene expression system for drug-controllable expression of transgenes and small hairpin RNAs (shRNAs). This system is highly robust and versatile, governing tightly controlled expression of transgenes and endogenous cellular genes (through shRNAs) in various primary and established cell lines in vitro, as well as in vivo in the central nervous system or in human cancer cells xenotransplanted into nude mice. The goal of this article is to provide a concise methodology for construction and manipulation of this conditional lentiviral-based system, and quantitative analyses of drug-inducible transgene expression and gene knockdown both in vitro and in vivo.
能够控制哺乳动物细胞中转基因表达和基因敲低的药物诱导系统是遗传研究中非常宝贵的工具，在转化研究或基因治疗中也可能发挥重要作用。我们和其他研究人员已经开发出一种基于慢病毒载体的条件性基因表达系统，用于转基因和小发夹RNA（shRNA）的药物可控表达。该系统高度稳健且用途广泛，可在体外多种原代细胞系和已建立的细胞系中，以及在中枢神经系统或异种移植到裸鼠体内的人类癌细胞中，严格控制转基因和内源性细胞基因（通过shRNA）的表达。本文的目的是提供一种简洁的方法，用于构建和操作这种基于条件性慢病毒的系统，以及对体外和体内药物诱导的转基因表达和基因敲低进行定量分析。
