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Efficient mouse transgenesis using Gateway-compatible ROSA26 locus targeting vectors and F1 hybrid ES cells

By Omar Nyabi, Michael Naessens, Katharina Haigh, Agnieszka Gembarska, Steven Goossens, Marion Maetens, Sarah De Clercq, Benjamin Drogat, Lieven Haenebalcke, Sonia Bartunkova, Ilse De Vos, Bram De Craene, Mansour Karimi, Geert Berx, Andras Nagy, Pierre Hilson, Jean-Christophe Marine and Jody J. Haigh

Abstract

The ability to rapidly and efficiently generate reliable Cre/loxP conditional transgenic mice would greatly complement global high-throughput gene targeting initiatives aimed at identifying gene function in the mouse. We report here the generation of Cre/loxP conditional ROSA26-targeted ES cells within 3–4 weeks by using Gateway® cloning to build the target vectors. The cDNA of the gene of interest can be expressed either directly by the ROSA26 promoter providing a moderate level of expression or by a CAGG promoter placed in the ROSA26 locus providing higher transgene expression. Utilization of F1 hybrid ES cells with exceptional developmental potential allows the production of germ line transmitting, fully or highly ES cell-derived mice by aggregation of cells with diploid embryos. The presented streamlined procedures accelerate the examination of phenotypical consequences of transgene expression. It also provides a unique tool for comparing the biological activity of polymorphic or splice variants of a gene, or products of different genes functioning in the same or parallel pathways in an overlapping manner

Topics: Methods Online
Publisher: Oxford University Press
OAI identifier: oai:pubmedcentral.nih.gov:2673446
Provided by: PubMed Central
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