Chapter title |
Establishment of Time- and Cell-Specific RNAi in Caenorhabditis elegans.
|
---|---|
Chapter number | 6 |
Book title |
Eukaryotic Transcriptional and Post-Transcriptional Gene Expression Regulation
|
Published in |
Methods in molecular biology, January 2017
|
DOI | 10.1007/978-1-4939-6518-2_6 |
Pubmed ID | |
Book ISBNs |
978-1-4939-6516-8, 978-1-4939-6518-2
|
Authors |
Masayuki Hamakawa, Takaaki Hirotsu |
Editors |
Narendra Wajapeyee, Romi Gupta |
Abstract |
The nematode worm Caenorhabditis elegans, in which loss-of-function mutants and RNA interference (RNAi) models are available, is a model organism useful for analyzing effects of genes on various life phenomena. In particular, RNAi is a powerful tool that enables time- or cell-specific knockdown via heat shock-inducible RNAi or cell-specific RNAi. However, the conventional RNAi methods are insufficient for investigating pleiotropic genes with various sites of action and life stage-dependent functions. To investigate the temporal- and cell-specific profiles of multifunctional genes, we established a new RNAi method that enables simultaneous time- and cell-specific knockdown (T.C.RNAi) in C. elegans. In this method, one RNA strand is expressed by a cell-specific promoter and the other by a heat shock promoter, resulting in only expression of double-stranded RNA in the target cell when heat shock is induced. We confirmed the effect of T.C.RNAi by the knockdown of GFP and the odr-3 gene which encodes Gα and is essential for olfaction. Further, this technique revealed that the control of glutamate receptors GLR-1 localization in RMD motor neurons requires Ras at the adult stage to regulate locomotion behavior. |
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