Fig. 4. Generation of GFR3 knockout mice by homologous recombination. (A) Gene targeting vector. A Neo cassette replaced a fragment containing exon one, deleting the start codon and the signal sequence of GFR3. H, HindIII; E, EcoRI; X, XhoI. (B) Southern blot screening of targeted ES clones. Genomic DNA was digested with HindIII. The probe, upstream of the short arm (shown in A), detected the 11 kb wild type and 7 kb mutant fragments. (C) PCR genotyping of tail DNA prepared from offspring of intercrossing heterozygous mice. Tail DNA was amplified with a primer set specific for the Neo gene, which detects the mutant allele, and a primer set that hybridises within the sequence that is deleted in the mutant and therefore detects the wild-type allele.