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Nucleic Acids Research 2005 33(7):e67; doi:10.1093/nar/gni065
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Published online 14 April 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oupjournals.org

Methods Online

Single copy shRNA configuration for ubiquitous gene knockdown in mice

Jost Seibler1,*, Birgit Küter-Luks1, Heidrun Kern1, Sandra Streu1, Leona Plum2,3, Jan Mauer2, Ralf Kühn1, Jens C. Brüning2 and Frieder Schwenk1,4

1Artemis Pharmaceuticals GmbH Neurather Ring 1, 51063 Cologne, Germany 2Department of Mouse Genetics and Metabolism and Center for Molecular Medicine (CMMC), Institute for Genetics Germany 3Klinik II and Poliklinik für Innere Medizin, University of Cologne Germany 4Department of Applied Natural Sciences, University of Applied Science Gelsenkirchen August-Schmidt-Ring 10, 45665 Recklinghausen, Germany

*To whom correspondence should be addressed. Tel: +49 221 9645342; Fax: +49 221 964 5321; Email: j.seibler{at}artemispharma.de

Received January 24, 2005. Revised March 18, 2005. Accepted March 18, 2005.

RNA interference through the expression of small hairpin RNA (shRNA) molecules has become a very promising tool in reverse mouse genetics as it may allow inexpensive and rapid gene function analysis in vivo. However, the prerequisites for ubiquitous and reproducible shRNA expression are not well defined. Here we show that a single copy shRNA-transgene can mediate body-wide gene silencing in mice when inserted in a defined locus of the genome. The most commonly used promoters for shRNA expression, H1 and U6, showed a comparably broad activity in this configuration. Taken together, the results define a novel approach for efficient interference with expression of defined genes in vivo. Moreover, we provide a rapid strategy for the production of gene knockdown mice combining recombinase mediated cassette exchange and tetraploid blastocyst complementation approaches.

Present address: Ralf Kühn, GSF Research Center, Institute for Developmental Genetics, Neuherberg/Munich, Germany


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