We have studied the linkage between the binding of Polycomb Group (PcG) proteins and the developmental regulation of gene expression using whole-genome mapping to identify sites bound by the PcG proteins, Polycomb (Pc) and Pleiohomeotic (Pho), in the Drosophila embryo and in a more restricted tissue, the imaginal discs of the third thoracic segment. Our data provide considerable support for the idea that Pho is a general component of the maintenance machinery since the majority of Pc targets are also associated with Pho binding. We find, in general, considerable developmental stability of Pc and Pho binding at target genes and observe that Pc/Pho binding can be associated with both expressed and inactive genes. In particular, at the Hox complexes both active and inactive genes have significant Pc and Pho binding; however, in comparison to inactive genes the active Hox genes show reduced and altered binding profiles. Keywords: ChIP-on-chip, tissue-specific binding Overall design: We used chromatin from whole embryos and larval T3 imaginal discs (haltere and third leg) to compare genomic binding of Pc and Pho between these tissues by ChIP-on-chip. Each chromatin source was represented with at least three biological replicates (in addition, Pc embryonic material and controls were hybridised to up to three different arrays), and enrichment profiles were generated by comparison of specific and control ChIP DNA samples. For Pc target analysis the specific reaction used chromatin from the Pc-GFP fly line immunopurified using anti-GFP, and the control reaction used wild type chromatin immunopurified using anti-GFP. For Pho analysis wild type chromatin was used with anti-Pho for the specific reaction and pre-immune antiserum for the control. Purified DNAs were then fragmented, TdT labeled and hybridized to the Affymetrix Drosophila genome Tiling Array 1.0 (reverse part number 520,054), as previously described (Manak et al. 2006).