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Nucleic Acids Research 2005 33(18):e155; doi:10.1093/nar/gni146
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Published online 12 October 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}oxfordjournals.org

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An ES cell system for rapid, spatial and temporal analysis of gene function in vitro and in vivo

Junhao Mao, Jeffery Barrow1, Jill McMahon, Joe Vaughan and Andrew P. McMahon*

Department of Molecular and Cellular Biology, Harvard University 16 Divinity Avenue, Cambridge, MA 02138, USA 1Department of Physiology and Developmental Biology, Brigham Young University 355 WIDB, Provo, UT 84602, USA

*To whom correspondence should be addressed. Tel: +1 617 496 3757; Fax: +1 617 496 3763; Email: amcmahon{at}mcb.harvard.edu

Received August 17, 2005. Revised September 14, 2005. Accepted September 14, 2005.

We describe a versatile genetic system for rapid analysis of mammalian gene function. In this, loss of reporter activity in a novel embryonic stem (ES) cell line enables rapid identification of targeting to the ubiquitously expressed Rosa26 locus. Subsequent regulation of gene activity is governed by a dual regulatory strategy utilizing two drugs, Tamoxifen and Doxycycline. To illustrate this approach, a dominant allele of Smoothened was introduced into this cell line, enabling regulated activation of Hedgehog signaling. By coupling Cre-loxP dependent activation with tetracycline dependent transcription in a single allele, we established a conditional method to control Smoothened activity and neural progenitor specification in differentiating ES cells in vitro and in chimeric embryos in vivo When crossed to an appropriate Cre driver strain, gene activity can also be temporally regulated within a specific cell lineage. This platform will facilitate rapid analysis of gene function in the mouse.


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