(A) bcr-targeting constructs. The structure of the bcr gene in the proximity of exon 1, the bcr-ABL targeting replacement vector and the predicted structure of the targeted bcr locus after homologous recombination, the bcr targeting vector, and the predicted structure of the bcr targeted allele. Only the relevant restriction sites are shown. B, BamHI; Xh, Xho I; R, EcoRI. Horizontal single arrows indicate the direction of transcription of the MC1-neo-pA and PGK-hyg-pA cassettes. Expected lengths of restriction fragments diagnostic for homologous recombination are indicated by double-headed arrows. The probe used to detect the homologous recombination events (BRA) in Southern filter hybridizations is indicated. (B) Expression of the bcr-ABL^p190 fusion gene in ES cells and in chimeric mouse tissues. mRNA was obtained from ES clones with bcr-ABL targeted alleles and from different chimeric mouse tissues. Reverse transcriptase was performed on cDNA made from each RNA preparation using primers specific for BCR-ABL cDNA. Products of the PCR reactions were separated on 1.2% agarose and hybridized with a specific internal oligo as a probe (5′-CCTTCAGCGGCCAGTAGCATCTGACTT-3′). ES-SA1 is a bcr-ABL/bcr⁺ ES clone and ES-SA0 is a bcr-ABL/bcr⁻ ES clone.