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  Generation of single and double knockdowns in polarized epithelial cells by retrovirus-mediated RNA interference.

Schuck, S., Manninen, A., Honsho, M., Fullekrug, J., & Simons, K. (2004). Generation of single and double knockdowns in polarized epithelial cells by retrovirus-mediated RNA interference. Proceedings of the National Academy of Sciences, USA, 101(14), 4912-4917.

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 Creators:
Schuck, Sebastian1, Author           
Manninen, Aki1, Author           
Honsho, Masanori1, Author           
Fullekrug, Joachim1, Author           
Simons, Kai1, Author           
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1Max Planck Institute of Molecular Cell Biology and Genetics, Max Planck Society, ou_2340692              

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 Abstract: RNA interference (RNAi) is a ubiquitous mechanism of eukaryotic gene regulation that can be exploited for specific gene silencing. Retroviruses have been successfully used for stable expression of short hairpin RNAs in mammalian cells, leading to persistent inhibition of gene expression by RNAi. Here, we apply retrovirus-mediated RNAi to epithelial Madin-Darby canine kidney cells, whose properties limit the applicability of other RNAi methods. We demonstrate efficient suppression of a set of 13 target genes by retroviral coexpression of short hairpin RNAs and a selectable marker. We characterize the resulting knockdown cell populations with regard to composition and stability, and examine the usefulness of proposed guidelines for choosing RNAi target sequences. Finally, we show that this system can be used to simultaneously target two genes, giving rise to double knockdowns. Thus, retrovirus-mediated RNAi is a convenient method for gene silencing in Madin-Darby canine kidney cells, and is likely to be applicable to virtually any mammalian cell.

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 Dates: 2004
 Publication Status: Issued
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 Identifiers: eDoc: 229774
Other: 369
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Title: Proceedings of the National Academy of Sciences, USA
Source Genre: Journal
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Pages: - Volume / Issue: 101 (14) Sequence Number: - Start / End Page: 4912 - 4917 Identifier: -