Chromosome 9q22 is a gene-rich region to which several human disease loci have been mapped. Pulsed field gel electrophoresis (PFGE) and FISH were used to determine the order of and distance between 12 chromosome 9q22 markers flanked by D9S196 and D9S180. D9S780 and XPA were within 190 kb of each other and hybridized to the same 460-kb NotI fragment as D9S180. ZNF169, a novel kruppel-type gene, and D9S280 shared several PFGE fragments indicating that they are not more than 300 kb apart. Interphase FISH showed that COL15A1 lies distal to the region bounded by D9S180 and D9S196 and that ZNF169 is adjacent to D9S196. Based on the restriction fragment lengths in this region and estimates from FISH, the distance from D9S180 to D9S196 is not less than 2 Mb.