Fig. 1. Notch1 and the transgenic constructs. Notch1 is a type-I transmembrane protein with a cleavable signal peptide followed by 36 EGF-like repeats and three Lin/Notch domains (LN). The intracellular domain contains ankyrin-like repeats, a polyglutamine stretch (Opa) and a protein stability PEST sequence. The first coding exon of the Notch1 gene was flanked by LoxP sequences (triangles) to generate the Floxed Notch1 allele (1) and the null allele for Notch1 generated after Cre-induced recombination (2). EcoRI (RI) restriction sites flanking the Floxed coding exon are shown. The Cre-recombinase was driven from the engrailed-2 promoter enhancer (En2-Cre), which restricted expression to the neuroepithelium of the midbrain-hindbrain boundary (Zinyk et al., 1998). The ROSA26-R Cre-reporter allele (R26R) was used in combination with the En2-Cre transgene and Floxed Notch1 alleles for cell fate tracing. The R26R comprises a Floxed PGK promoter driving a neomycin cassette containing four polyadenylation sequences (4XpA) and a ß-galactosidase coding region (lacZ) with polyadenylation site (bpA) introduced into the ROSA26 locus by homologous recombination (Soriano, 1999). Cre-mediated recombination deletes the PGK neo cassette and results in constitutive expression of ß-galactosidase.
