There is growing evidence that the histone-DNA complexes found in nucleosomes offer protection from DNA degradation processes, including apoptotic events in addition to bacterial and environmental degradation. We sought to locate human nucleosome regions and build a catalogue of SNPs sited near the middle of these genomic segments that could be combined into a single PCR multiplex specifically for use with extremely degraded human genomic DNA samples. Using recently optimized bio-informatics tools for the reliable identification of nucleosome sites based on sequence motifs and their positions relative to known promoters, 1395 candidate loci were collected to construct an 18-plex single base extension assay. Genotyping performance of the nucleosome SNPs was tested using artificially degraded DNA and 24 casework samples where the likely state of degradation of DNA was established by comparison to profile completeness in four other forensic assays: a standard 15-plex STR identification test, a miniaturized STR multiplex and two autosomal SNP multiplexes. The nucleosome SNP assay gave genotyping success rates 6% higher than the best existing forensic SNP assay: the SNPforID Auto-2 29-plex and significantly higher than the mini-STR assay. The nucleosome SNPs we located and combined therefore provide a new type of marker set that can be used to supplement existing approaches when the analysed DNA is likely to be extremely degraded and may fail to give sufficient STR genotypes for a reliable identification.