We have generated a mouse line, MORE (Mox2Cre), which expresses the Cre recombinase from the Mox2 locus. This locus drives expression of Cre throughout the epiblast following implantation. A nls-Cre recombinase gene (provided by Steve O’Gorman) was inserted by homologous recombination in embryonic stem (ES) cells at the initiator ATG of the Mox2 gene, which maps to mouse chromosome 12 (Candia et al., 1992). The Cre gene was followed immediately by a neomycin resistance cassette (Fig. 1a). To maintain fidelity of expression from the Mox2 locus, we replaced only the first coding exon and 300 bp of the following intron. Correct recombination events in ES cells were confirmed by Southern blot (Fig. 1b and c). The insertion should create a null allele for Mox2 because it replaces the same exon as a recently reported Mox2 disruption, and the MORE mice exhibit a phenotype identical to that which was described (Mankoo et al. 1999). Homozygous mice are viable, but some die before weaning. The surviving mice have defects in limb musculature resulting in an abnormal limb posture.

Based on several reports of Mox2 RNA expression (Candia and Wright, 1996; Candia et al., 1992; Mankoo et al., 1999), we had anticipated that Cre from the Mox2 locus would be expressed in the early epithelial somite and then become restricted to the sclerotome and limb musculature. Instead, we found that Cre activity from the Mox2 locus is detected as early as E5. To follow the Cre activity, we mated MORE mice to R26R mice (Soriano, 1999). Figure 2 illustrates the floxed R26R activity at different stages throughout embryogenesis. No β-galactosidase positive cells were found in E3. 5-day-old embryos (data not shown), but a number of β-galactosidase positive cells were present in inner cell mass (ICM) outgrowths from explanted blastocysts 2 days after hatching (Fig. 2a). In vivo, the first detectable expression of Cre is at E5. Figure 2b