Analysis of gene function in somatic mammalian cells using small interfering RNAs

• Published in: Methods (San Diego, Calif.) Vol. 26; no. 2; pp. 199 - 213
• Main Authors: Elbashir, Sayda M., Harborth, Jens, Weber, Klaus, Tuschl, Thomas
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.02.2002
• Subjects: Animals; Base Sequence; Blotting, Western; Cell Line; Cytoskeleton; DNA; Dose-Response Relationship, Drug; Double-stranded RNA; Drosophila; Gene Silencing; Genes, Reporter; Genetic Techniques; HeLa Cells; Humans; Knockdown; Luciferases - metabolism; Microscopy, Fluorescence; Models, Genetic; Molecular Sequence Data; Oligonucleotides - chemistry; Posttranscriptional gene silencing; RNA - chemistry; RNA - metabolism; RNA interference; RNA, Messenger - metabolism; RNA, Small Interfering; RNA, Untranslated - chemistry; RNA, Untranslated - genetics; Small interfering RNA; Transfection; Small interfering RNA; Knockdown; RNA interference; Posttranscriptional gene silencing; Double-stranded RNA
• ISSN: 1046-2023; 1095-9130
• DOI: 10.1016/S1046-2023(02)00023-3

RNA interference (RNAi) is a highly conserved gene silencing mechanism that uses double-stranded RNA (dsRNA) as a signal to trigger the degradation of homologous mRNA. The mediators of sequence-specific mRNA degradation are 21- to 23-nt small interfering RNAs (siRNAs) generated by ribonuclease III cleavage from longer dsRNAs. Twenty-one-nucleotide siRNA duplexes trigger specific gene silencing in mammalian somatic cells without activation of the unspecific interferon response. Here we provide a collection of protocols for siRNA-mediated knockdown of mammalian gene expression. Because of the robustness of the siRNA knockdown technology, genomewide analysis of human gene function in cultured cells has now become possible.