Paddison, P. J., Caudy, A. A., Sachidanandam, R., Hannon, G. J. (2004) Short hairpin activated gene silencing in mammalian cells. In: RNA Interference, Editing and Modification: methods and protocols. Methods Mol Biol, 265 . Springer, pp. 85-100.
Abstract
RNA interference (RNAi) is now a popular method for silencing gene expression in a variety of systems. RNAi methods use double-stranded RNAs (dsRNAs) to target complementary RNAs for destruction. In mammalian systems, very short dsRNAs (22-25 bp) such as short interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) are used to avoid endogenous nonspecific antiviral responses that target longer dsRNAs. siRNAs elicit a transient silencing response, while shRNAs can be expressed continuously to establish stable gene silencing. shRNAs can be introduced into cells and animals using a variety of standard vectors as well as retroviral or lentiviral expression systems. This chapter describes the design, construction, validation, and use of shRNAs for silencing genes. We report our results from testing a variety of shRNA design features and shRNA expression vectors. We also provide methods that use shRNAs to permit different levels of gene expression. Additionally, we discuss some aspects important for constructing an information pipeline to support development of a large shRNA library.
| Item Type: | Book Section |
|---|---|
| Uncontrolled Keywords: | Animals Base Sequence Cloning Molecular Gene Silencing physiology Humans Mammals Molecular Sequence Data Nucleic Acid Conformation Plasmids Polymerase Chain Reaction methods RNA Interference physiology RNA Small Nuclear genetics Restriction Mapping Reverse Transcriptase Polymerase Chain Reaction methods |
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > RNAi bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > genes, structure and function > gene silencing bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > siRNA |
| CSHL Authors: | |
| Communities: | CSHL labs > Hannon lab School of Biological Sciences > Publications |
| Depositing User: | CSHL Librarian |
| Date: | 2004 |
| Date Deposited: | 26 Jan 2012 16:32 |
| Last Modified: | 19 Sep 2014 15:07 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/22454 |
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