Comparative cytogenetic analysis of the genomes of the model grass Brachypodium distachyon and its close relatives

Brachypodium is a small genus of temperate grasses that comprises 12-15 species. Brachypodium distachyon is now well established as a model species for temperate cereals and forage grasses. In contrast to B. distachyon, other members of the genus have been poorly investigated at the chromosome level or not at all. Twenty accessions comprising six species and two subspecies of Brachypodium were analysed cytogenetically. Measurements of nuclear genome size were made by flow cytometry. Chromosomal localization of 18-5 center dot 8-25S rDNA and 5S rDNA loci was performed by dual-colour fluorescence in situ hybridization (FISH) on enzymatically digested root-tip meristematic cells. For comparative phylogenetic analyses genomic in situ hybridization (GISH) applied to somatic chromosome preparations was used. All Brachypodium species examined have rather small genomes and chromosomes. Their chromosome numbers and genome sizes vary from 2n = 10 and 0 center dot 631 pg/2C in B. distachyon to 2n = 38 and 2 center dot 57 pg/2C in B. retusum, respectively. Genotypes with 18 and 28 chromosomes were found among B. pinnatum accessions. GISH analysis revealed that B. pinnatum with 28 chromosomes is most likely an interspecific hybrid between B. distachyon (2n = 10) and B. pinnatum (2n = 18). Two other species, B. phoenicoides and B. retusum, are also allopolyploids and B. distachyon or a close relative seems to be one of their putative ancestral species. In chromosomes of all species examined the 45S rDNA loci are distally distributed whereas loci for 5S rDNA are pericentromeric. The increasing significance of B. distachyon as a model grass emphasizes the need to understand the evolutionary relationships in the genus Brachypodium and to ensure consistency in the biological nomenclature of its species. Modern molecular cytogenetic techniques such as FISH and GISH are suitable for comparative phylogenetic analyses and may provide informative chromosome- and/or genome-specific landmarks.