AbstractEleven site‐specific gene probes derived from human chromosome 21 were assigned to each region on the original chromosome byin situhybridization. These probes were isolated from a lambda library, which contains inserts from chromosome 21 in the human‐mouse hybrid cell WA17. This cell contains chromosome 21 as its only human chromosome. These single copy inserts had unique DNA sequence and had been screened for restriction fragment length polymorphism (RFLP) in the DNA of unrelated humans.Among the eleven probes, five were assigned to 21q22.3, two to 21 ql 1, and the four remaining genes were respectively assigned to 21 ql 1 ‐ 21q21, 21q21, 21q21 ‐ 21q22.1, and a centromeric region. The gene locus of superoxide dismutase‐1 (SOD‐1) was determined to be at 21q22.1. Since region q22 is suggested to be the gene loci of the main features of Down syndrome, probes in the region may yield information about this disorder. A pericentric probe is useful in determining predisposition to non‐disjunction in 21 trisomy. We determined gene loci for these site‐specific gene probes in order to make a complete gene map of