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J. Biol. Chem., Vol. 283, Issue 9, 5899-5907, February 29, 2008

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 283/9/5899    most recent M709410200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via Google Scholar Google Scholar Articles by 't Hoen, P. A. C. Articles by den Dunnen, J. T. Search for Related Content PubMed PubMed Citation Articles by 't Hoen, P. A. C. Articles by den Dunnen, J. T. Social Bookmarking                      What's this?

Generation and Characterization of Transgenic Mice with the Full-length Human DMD Gene^*

Peter A. C. 't Hoen¹, Emile J. de Meijer, Judith M. Boer, Rolf H. A. M. Vossen, Rolf Turk, Ronald G. H. J. Maatman, Kay E. Davies, Gert-Jan B. van Ommen, Judith C. T. van Deutekom, and Johan T. den Dunnen

From the Center for Human and Clinical Genetics, Leiden University Medical Center, Postal Zone S4-P, P.O. Box 9600, 2300 RC Leiden, The Netherlands and the Department of Physiology, Anatomy, and Genetics, Medical Research Council Functional Genetics Unit, Oxford, OX1 3QX United Kingdom

We report the generation of mice with an intact and functional copy of the 2.3-megabase human dystrophin gene (hDMD), the largest functional stretch of human DNA thus far integrated into a mouse chromosome. Yeast spheroplasts containing an artificial chromosome with the full-length hDMD gene were fused with mouse embryonic stem cells and were subsequently injected into mouse blastocysts to produce transgenic hDMD mice. Human-specific PCR, Southern blotting, and fluorescent in situ hybridization techniques demonstrated the intactness and stable chromosomal integration of the hDMD gene on mouse chromosome 5. Expression of the transgene was confirmed by RT-PCR and Western blotting. The tissue-specific expression pattern of the different DMD transcripts was maintained. However, the human Dp427p and Dp427m transcripts were expressed at 2-fold higher levels and human Dp427c and Dp260 transcripts were expressed at 2- and 4-fold lower levels than their endogenous counterparts. Ultimate functional proof of the hDMD transgene was obtained by crossing of hDMD mice with dystrophin-deficient mdx mice and dystrophin and utrophin-deficient mdx x Utrn^-/- mice. The hDMD transgene rescued the lethal dystrophic phenotype of the mdx x Utrn^-/- mice. All signs of muscular dystrophy disappeared in the rescued mice, as demonstrated by histological staining of muscle sections and gene expression profiling experiments. Currently, hDMD mice are extensively used for preclinical testing of sequence-specific therapeutics for the treatment of Duchenne muscular dystrophy. In addition, the hDMD mouse can be used to study the influence of the genomic context on deletion and recombination frequencies, genome stability, and gene expression regulation.

Received for publication, November 15, 2007

^* This work was financially supported by Muscular Dystrophy Association (United States) Project 3562, the Muscular Dystrophy Campaign (United Kingdom) Project RA3/647, and the Center for Biomedical Genetics. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. 1 and 2 and Tables 1 and 2.

¹ Recipient of a VENI grant from the Dutch Organization for Scientific Research (NWO Grant 2005/03808/ALW). To whom correspondence should be addressed. Tel.: 31-71-5269421; Fax: 31-71-5268285; E-mail: p.a.c.hoen{at}lumc.nl.

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