The economic and ecological importance of forest trees, as well as their unique biological features, has recently raised the level of interest in studies on their genomes, including sequencing of the entire poplar genome. However, cytogenetic studies have not moved in parallel with developments in genomics. This is especially true for hardwood species characterized by small genomes and relatively high numbers of small chromosomes. Molecular cytogenetic studies have mainly been focused on coniferous species, owing to the larger size of their chromosomes, and have been applied exclusively for chromosome identification and comparative karyotyping in an attempt to understand genome evolution and phylogenetic relationships. In this context, rRNA genes physical mapped by FISH reveal particularly useful chromosomal landmarks with variable distribution patterns between species. Here we present a contribution of DNA markers used for chromosome analysis, which already allowed a deeper characterization and understanding of the processes underlying genome diversity of forest trees. The use of advanced cytogenetic techniques and other potential important methods for genome analysis of forest trees is also discussed.

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