Janas, J. A., Skowronski, J., Van Aelst, L. (2006) Lentiviral delivery of RNAi in hippocampal neurons. Methods Enzymol, 406. pp. 593-605. ISSN 0076-6879 (Print)
Abstract
The breakthrough discovery that double-stranded RNA of 21 nucleotides in length (referred to as short or small interfering RNA; siRNA) can trigger sequence-specific gene silencing in mammalian cells has led to the development of a powerful new approach to study gene function (Dillon et al., 2005; Dykxhoorn et al., 2003; Elbashir et al., 2001; Hannon et al., 2004). Effective delivery of siRNA molecules into target cells or tissues is critical for successful RNA interference (RNAi) application. Here, we describe the use of human immunodeficiency virus type 1 (HIV-1)-based lentiviral vectors for delivery of short hairpin RNA (shRNA), a precursor of siRNA, into primary neurons to suppress gene expression. Major advantages of lentiviral vectors are their ability to transduce nondividing cells and to confer long-term expression of transgenes. This chapter covers selection of short hairpin sequences, vector design, production of lentiviral supernatants, transduction of dissociated primary hippocampal neurons, and testing the effectiveness of shRNA-mediated silencing.
| Item Type: | Paper |
|---|---|
| Uncontrolled Keywords: | Animals Cloning Molecular Female Gene Silencing Genetic Vectors Hippocampus cytology embryology Lentivirus Neurons drug effects RNA Interference physiology RNA Small Interfering pharmacology Rats |
| Subjects: | diseases & disorders > viral diseases > HIV bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > RNAi organs, tissues, organelles, cell types and functions > tissues types and functions > hippocampus organism description > virus > lentivirus bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > siRNA |
| CSHL Authors: | |
| Communities: | CSHL labs > Van Aelst lab CSHL Post Doctoral Fellows |
| Depositing User: | CSHL Librarian |
| Date: | 2006 |
| Date Deposited: | 14 Dec 2011 15:51 |
| Last Modified: | 02 May 2013 18:36 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/22829 |
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