The gene was disrupted by replacement of exon 3 and part of exon 4 (encoding amino acids 92-154) with a PGK-neo cassette via homologous recombination. Introduction of PGK-neo introduces stop codons into all 3 reading frames. Western blot analysis of brain lysates from homozygous mutant animals using antibodies directed against the C-terminus of the protein confirmed the absence of protein expression. Immunoprecipitation assays also confirmed the absence of protein product. Northern blot and RT-PCR analysis detected mutant transcripts containing neo and lacking exon 3 and part of exon 4 in brain and skeletal muscles from homozygous mutants.