High-throughput RNAi screening in cultured cells: a user's guide

• Published in: Nature reviews. Genetics Vol. 7; no. 5; pp. 373 - 384
• Main Authors: Echeverri, Christophe J, Perrimon, Norbert
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group 01.05.2006
• Subjects: Animals; Automation; Biological and medical sciences; Cells; Cells, Cultured; Drosophila melanogaster - genetics; Fundamental and applied biological sciences. Psychology; Gene Expression Profiling - methods; Gene Transfer Techniques; Genetics of eukaryotes. Biological and molecular evolution; Genomes; Humans; Insects; Interferon; Models, Biological; Optimization; Phenotype; Physiology; Quality Control; Reagents; RNA Interference; RNA, Small Interfering - genetics; Signal transduction; Screening; Gene silencing; RNA interference
• ISSN: 1471-0056; 1471-0064
• DOI: 10.1038/nrg1836

RNA interference has re-energized the field of functional genomics by enabling genome-scale loss-of-function screens in cultured cells. Looking back on the lessons that have been learned from the first wave of technology developments and applications in this exciting field, we provide both a user's guide for newcomers to the field and a detailed examination of some more complex issues, particularly concerning optimization and quality control, for more advanced users. From a discussion of cell lines, screening paradigms, reagent types and read-out methodologies, we explore in particular the complexities of designing optimal controls and normalization strategies for these challenging but extremely powerful studies.