Fig. 1. Schematic representation of the Mgn^tZ targeting vector and recombination at the Mgn locus. (A) Restriction map of the wild-type Mgn locus. Broken lines indicate regions of homology in the targeting vector. (B) Restriction map of the Mgn^tZ targeting vector. (C) The predicted structure of a mutated Mgn allele following homologous recombination. The horizontal bars (5' and 3' external probe) indicate the DNA fragment used for Southern blot analysis. (D) Targeted locus after removing neomycin cassette (neo) by Cre-mediated excision at the loxP sites. (E) Southern blot analysis of restriction enzyme-digested DNA from targeted Mgn^tZ (+/tZ) and wild-type (+/+) animals with 5' and 3' external probes (the same strategy was used to detect littermates from heterozygous intercrosses). wt and mt indicate the position of the wild-type and the mutated allele, respectively. (F) Southern blot containing genomic DNA upon SacII digestion and probed with an internal probe, indicating proper excision of the neomycin gene (neo). (G) Heteroduplex PCR (primers for the mutant allele, MM107 and tlacZ; primers for the wild-type allele, MM108 and MM109) used to identify mutated embryos/mice. (H) RT-PCR with specific primers for the bHLH domain of Mgn (MM111D and MM112R). The 250 bp band is missing in the Mgn^tZ/tZ mice.