A FRT-loxP-neo-FRT-LoxP cassette was inserted upstream exon 2 and a single loxP site was inserted between exon 5 and 6. The frt-flanked neomycin cassette was removed by FLP-mediated recombination, followed by cre-mediated recombination in the germline, removing exons 2-5. Western blot analysis confirmed the complete absence of protein expression in primary cultures of hippocampal neurons derived from E18 homozygous mutant mice.