Clonal analysis in the intact mouse embryo by intragenic homologous recombination.
A method of cell labelling based on homologous recombination has been developed to analyze the lineage of cells in intact embryos. In this method a LacZ gene bearing an inactivating duplication in its coding frame (termed LaacZ gene) is inserted into the genome of mice. Subsequently, in transgenic embryos, homologous recombination within the duplication recreates an open reading frame for beta-galactosidase. The descendants of the modified cell are identified histochemically. This recombination occurs at random and with low frequency. It can occur in cells in which the gene is not transcribed. It can also be intragenic (in adjacent sequences) as single transgene copies are found to recombine. To analyze the lineage of cells in the myotome the expression of the LaacZ gene is driven by a promoter which confers expression specifically to cells of this compartment. The analysis in 40-somite embryos (E11.5) is reported. We show that individual beta-gal+ myotomal clones may contribute to somites from both sides of the animal, in which case the right and the left somites are labelled consecutively and at the same antero-posterior level. It reflects predictable clonal origin of myotomal cells at the primitive streak stage or before and demonstrates three morphogenetic movements in the mesoderm: bilateral, antero-posterior and mediolateral. The results also show that the left-right assignation of myotomal cells in mouse is late, probably occurring during cell migrations at gastrulation. It applies also to presumptive myotomal cells of limb musculature. These results contrast with early left-right assignation of most tissues in Xenopus and Zebrafish. Some clones contribute to distant somites, suggesting persistence of presumptive myotomal cells in the presumptive paraxial mesoderm and clonal organisation of the myotome.
Bonnerot
C
C
Unité de Biologie moléculaire du Développement, Institut Pasteur, Paris, France.
Nicolas
J F
JF
D000818:Animals; D015139:Blotting, Southern; D002478:Cells, Cultured; D003001:Cloning, Molecular / Q000379:methods*; D007763:Lac Operon / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic / Q000196:embryology*; D009132:Muscles / Q000166:cytology / Q000196:embryology; D011995:Recombination, Genetic; D001616:beta-Galactosidase / Q000235:genetics
D001616:beta-Galactosidase