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 m ethod a LacZ gene bearing an inactivating duplication in its coding fr ame (termed LaacZ gene) is inserted into the genome of mice. Subsequen tly, in transgenic embryos, homologous recombination within the duplic ation recreates an open reading frame for beta-galactosidase. The desc endants bf the modified cell are identified histochemically. This reco mbination occurs at random and with low frequency. It can occur in cel ls in which the gene is not transcribed. It can also be intragenic (in adjacent sequences) as single transgene copies arc found to recombine . To analyze the lineage of cells in the myotome the expression of the LaacZ gene is driven by a promotor which confers expression specifica lly to cells of this compartment. The analysis in 40-somite embryos (E 11.5) is reported. We show that individual beta-gal+ myotomal clones m ay contribute to somites from both sides of the animal, in which case the right and the left somites are labelled consecutively and at the s ame antero-posterior level. It reflects predictable clonal origin of m yotomal cells at the primitive streak stage or before and demonstrates three morphogenetic movements in the mesoderm : bilateral, antero-pos terior and mediolateral. The results also show that the left-right ass ignation of myotomal cells in mouse is late, probably occurring during cell migrations at gastrulation. It applies also to presumptive myoto mal cells of limb musculature. These results contrast with early left- right assignation of most tissues in Xenopus and Zebrafish. Some clone s contribute to distant somites, suggesting persistence of presumptive myotomal cells in the presumptive paraxial mesoderm and clonal organi sation of the myotome.