The YES1 proto-oncogene was mapped previously to human chromosome band 18q21.3 by using isotopic in situ hybridization. Using yeast artificial chromosomes (YACs) as probes and fluorescence in situ hybridization, a strong signal was detected in the region corresponding to 18p11.3. Restriction digests confirmed that the YACs contained the YES1 gene and not other cross-hybridizing, protein-tyrosine kinases. In addition, these-YACs were found to contain another 18p11.32 gene, thymidlylate synthase. These genes were less than 50 kb apart. Collectively, these data suggest that YES1 maps to 18p11.32 rather than to 18q21.3.