GENOME WIDE RNAI SCREENING AND ANALYSIS

点击翻译按钮获取中文摘要

Genetic screens have been a powerful approach for defining signaling and developmental pathways in model organisms, but historically have not been exploited in mammalian cancer biology because of a lack of systematic tools. Recent advances in RNA interference (RNAi) have begun to facilitate such approaches, and these genetic tools have become essential components of basic and translational cancer biology. The Genome-wide RNAi Screening and Analysis (GRSA) Shared Resource was established through philanthropic and institutional support in 2008 to facilitate investigators in their use of new RNAi technologies and genetic screening methods. The Shared Resource is directed by Drs. Thomas Westbrook and Dan Liu who have extensive expertise in developing genetic technologies (e.g. RNAi libraries) and in mammalian genetic screening methods. Combined with this expertise, the GRSA provides all essential elements for single-gene analyses to whole-genome genetic screens including genome-wide short-hairpin RNA (shRNA) libraries, multiple automated robotic platforms for library manipulation, high-throughput analyzers for phenotypic analysis, and data processing infrastructure for mammalian genetic screens, making this DLDCC Shared Resource unique among NCI Cancer Centers. Specific services provided by the Shared Resource include (1) performing whole-genome or sub-genome scale RNAi screens, (2) utilizing individual lentivirus-based shRNA vectors, (3) large-scale automated manipulation and preparation of shRNA libraries, (4) automated mammalian cell transfection and lentivirus production, (5) high-throughput cell-based assays using automated microscopy or flow cytometry, and (6) data analysis, storage, and management. By housing these resources in a single, cohesive Shared Resource, the GRSA enables investigators to employ genetic approaches that are often cost- and labor-prohibitive or simply not feasible for individual laboratories. In the current application, the GRSA is proposed as a new Shared Resource for the DLDCC grant application.