Hannon, G. J., Conklin, D. S. (2004) RNA interference by short hairpin RNAs expressed in vertebrate cells. Methods Mol Biol, 257. pp. 255-66. ISSN 1940-6029 (Electronic)1064-3745 (Linking)
Abstract
RNA interference (RNAi) is now established as a general method to silence gene expression in a variety of organisms. Double-stranded RNA (dsRNA), when introduced to cells, interferes with the expression of homologous genes, disrupting their normal function. In mammals, transient delivery of synthetic short interfering RNAs (siRNAs), which resemble the processed form of standard double stranded RNAi triggers, is effective in silencing mammalian genes. Issues related to transfer efficiency and duration of the silencing effect, however, restrict the spectrum of the applications of siRNAs in mammals. These shortcomings of siRNAs have been solved by the cellular expression of short hairpin RNAs (shRNAs) from DNA vectors. shRNAs are indistinguishable from siRNAs in terms of efficacy and mechanism but can be produced within cells from standard mammalian expression vectors. In this way, shRNA expression makes possible the creation of continuous cell lines and transgenic animals in which suppression of a target gene is stably maintained by RNAi. As a result, the types of RNAi-based gene function analysis that can be carried out in mammals have been greatly expanded. We describe methods for the construction and transfer of stable shRNA expressing vectors suitable for generating loss of function alleles in mammalian cells in vitro or in vivo.
| Item Type: | Paper |
|---|---|
| Uncontrolled Keywords: | Alleles Animals Cells Cultured Gene Expression Gene Silencing physiology Genetic Vectors Humans Mice RNA metabolism pharmacology RNA Interference RNA Polymerase III metabolism RNA Processing Post-Transcriptional physiology Ribonuclease III metabolism Transfection Vertebrates genetics metabolism |
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > RNAi |
| CSHL Authors: | |
| Communities: | CSHL labs > Hannon lab |
| Depositing User: | CSHL Librarian |
| Date: | 2004 |
| Date Deposited: | 09 Feb 2012 15:28 |
| Last Modified: | 01 Dec 2016 17:31 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/22379 |
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