Recently we demonstrated the clustering of deletion breakpoints in the pericentromeric region of human chromosome 17p in human primitive neuroectodermal tumors (PNETs). Chromosomal disruption was shown to occur between the two markers D17S805 and D17S953, a region previously shown to be deleted in the Smith-Magenis syndrome. To characterize the molecular basis of this genomic instability, we established clone contigs covering this region. An initial physical map of chromosome 17p has been constructed with overlapping sets of YACs. YAC clones were transformed into five clone contigs according to their content of 30 previously known and 16 newly established sequence-tagged sites (STSs). To circumvent the complications inherent in YAC technologies, such as internal deletions, chimerism, and complex rearrangements, we then converted the YAC contigs to PAC and cosmid contigs. Thirty-nine individual PAC/cosmid clones were identified and were used to construct six different PAC/cosmid contigs ranging from 130 to 1200 kb in size and covering approximately 2.5 Mb of genomic DNA. The composite YAC/PAC/cosmid map covers a region of > 6 Mb of genomic DNA consisting of four different clone contigs of up to 2.9 Mb in size. We have demonstrated that three STSs (D17S58, PS1, and D17S842) are duplicated, suggesting the occurrence of low abundant repetitive sequences in this region. By integration of publicly available information we further mapped 10 genes and ESTs to their precise chromosomal positions and thus could exclude or identify them as candidate genes for PNET and/or the Smith-Magenis syndrome.