FIG. 3.

(A) Targeting strategy for disruption of the mouse Neuralized locus. A targeting cassette containing an IRES-GFP construct and a PGK-neo cassette for positive selection of transfected clones was inserted into the exon encoding the NHR-2 region of the protein. A thymidine kinase cassette (PGK-TK) was used for negative selection with FIAU. (B) Genotyping was performed by Southern blot analysis using a 5′ external probe and genomic DNA digested with BamHI. The wild-type allele yields a 14-kbp fragment, and the mutant allele yields a 9-kbp band. The Southern blot shows a typical result from tail DNA of offspring from a heterozygous intercross and the appearance of all three expected genotypes. (C) PCR genotyping assay on tail DNA from offspring of a heterozygous intercross using a three-primer setup. The wild-type allele yields a PCR product of 367 bp, the mutant allele yields a product of 190 bp, and all three genotypes are readily detectable.