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Exon 12, encoding subdomains 5 and 6 of the catalytic domain, was replaced with a neomycin selection cassette. Northern blot analysis of total RNA isolated from ES cells identified the production of a novel transcript containing the neo transgene. RT-PCR analysis confirmed that transcription was derived from the endogenous promoter with read-through into the transgene. Western blot analysis failed to detect encoded protein in cell lysates from homozygous mutant mice. The functional ablation of the protein was determined via an immune complex kinase autophosphorylation assays. (J:53258)