A system for stable expression of short interfering RNAs in mammalian cells.
Mammalian genetic approaches to study gene function have been hampered by the lack of tools to generate stable loss-of-function phenotypes efficiently. We report here a new vector system, named pSUPER, which directs the synthesis of small interfering RNAs (siRNAs) in mammalian cells. We show that siRNA expression mediated by this vector causes efficient and specific down-regulation of gene expression, resulting in functional inactivation of the targeted genes. Stable expression of siRNAs using this vector mediates persistent suppression of gene expression, allowing the analysis of loss-of-function phenotypes that develop over longer periods of time. Therefore, the pSUPER vector constitutes a new and powerful system to analyze gene function in a variety of mammalian cell types.
Brummelkamp
Thijn R
TR
Division of Molecular Carcinogenesis, Division of Tumor Biology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, Netherlands.
Bernards
René
R
Agami
Reuven
R
D064173:Anaphase-Promoting Complex-Cyclosome; D018797:Cell Cycle Proteins; D015536:Down-Regulation; D020868:Gene Silencing; D016158:Genes, p53; D005821:Genetic Techniques; D005822:Genetic Vectors; D006801:Humans; D008025:Ligases / Q000235:genetics; D009154:Mutation; D009690:Nucleic Acid Conformation; D010641:Phenotype; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering; D022661:RNA, Untranslated / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes; D000097869:Polo-Like Kinase 1
D064173:Anaphase-Promoting Complex-Cyclosome; D018797:Cell Cycle Proteins; D008025:Ligases; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D022661:RNA, Untranslated; D016159:Tumor Suppressor Protein p53; D043743:Ubiquitin-Protein Ligase Complexes; D000097869:Polo-Like Kinase 1