A number of human telomeres have been successfully cloned using a modified yeast artificial chromosome (YAC) vector (half-YAC) cloning strategy, but to date, human chromosome 22q has not been identified by this approach. We used an alternative approach of genomic walking, starting from a subtelomeric sequence, Tel-Bam3.4. present on a number of human chromosomes including 22q. This approach was successful in the development of a cosmid contig representing the terminal 140 kb of human chromosome 22q, providing telomeric closure of the genetic and physical maps for 22q. The most distal region of the contig contains subtelomeric repeats which crosshybridize to a number of chromosomes, while the proximal sequences are unique for 22q. The unique sequence cosmid was used as a 22qter-specific probe for fluorescence in situ hybridization (FISH) analysis, which confirmed that this cosmid was distal to the most telomeric marker previously available for chromosome 22. In addition, this cosmid was used to document a 22q terminal deletion that was not detectable by conventional cytogenetic analysis. Unique telomere-specific FISH probes such as this one will have significant diagnostic value in the detection of cryptic deletions and translocations in patients with unexplained mental retardation and other patient populations.