The gene for tyrosine hydroxylase, the first and rate-limiting enzyme in the biosynthetic pathway of catecholamine neurotransmitters, has been localized in situ to chromosome 6 in the chicken. It is the first DNA marker to be reported for this telocentric macrochromosome. Use of a 45-kb biotinylated chicken-specific cosmid probe and a sensitive fluorescent detection system proved to be highly efficient, with over 90% of metaphases showing positive hybridization signals on one or (usually) both chromosome 6 homologs, in physically mapping this single-gene locus.