Selective disruption of genes transiently induced in differentiating mouse embryonic stem cells by using gene trap mutagenesis and site-specific recombination.
A strategy employing gene trap mutagenesis and site-specific recombination (Cre/loxP) has been used to identify genes that are transiently expressed during early mouse development. Embryonic stem cells expressing a reporter plasmid that codes for neomycin phosphotransferase and Escherichia coli LacZ were infected with a retroviral gene trap vector (U3Cre) carrying coding sequences for Cre recombinase (Cre) in the U3 region. Activation of Cre expression from integrations into active genes resulted in a permanent switching between the two selectable marker genes and consequently the expression of beta-galactosidase (beta-Gal). As a result, clones in which U3Cre had disrupted genes that were only transiently expressed could be selected. Moreover, U3Cre-activating cells acquired a cell autonomous marker that could be traced to cells and tissues of the developing embryo. Thus, when two of the clones with inducible U3Cre integrations were passaged in the germ line, they generated spatial patterns of beta-Gal expression.
Thorey
I S
IS
Department of Hematology, University of Frankfurt Medical School, Frankfurt am Main, Germany.
Muth
K
K
Russ
A P
AP
Otte
J
J
Reffelmann
A
A
von Melchner
H
H
D000818:Animals; D019521:Body Patterning / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics; D002999:Clone Cells; D005314:Embryonic and Fetal Development / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D019426:Integrases; D007763:Lac Operon; D051379:Mice / Q000235:genetics*; D008822:Mice, Transgenic; D008967:Molecular Biology / Q000379:methods*; D016296:Mutagenesis; D011995:Recombination, Genetic; D013234:Stem Cells; D014018:Tissue Distribution; D014764:Viral Proteins; D001616:beta-Galactosidase / Q000096:biosynthesis
D014764:Viral Proteins; C045073:Cre recombinase; D019426:Integrases; D001616:beta-Galactosidase