A neo cassette with a 3' loxP site was inserted downstream of exon 3 and an additional loxP site was inserted upstream of exon 3. Cre-mediated recombination in ES cells was used to remove the neo cassette and exon 3. The absence of protein product from the allele was confirmed by a 50% reduction in protein in heterozygotes as determined by western blot analysis on heart and kidney extracts.