A neomycin selection cassette replaced sequences encoding 47 amino acids of the Rel homology domain. An absence of normal protein in homozygous mutant mice was determined through Western blot analysis. Low levels of reactive polypeptides were present in the cytoplasm, however, no reactivity was observed in the nucleus. RT-PCR and sequence analysis revealed aberrantly spliced in-frame transcripts from which the targeted exon had been excised. An electrophoretic mobility shift assay confirmed that DNA was not binding to the mutant protein.