The Large-Scale Organization of the Centromeric Region in Beta Species

In higher eukaryotes, the DNA composition of centromeres displays a high degree of variation, even between chromosomes of a single species. However, the long-range organization of centromeric DNA apparently follows similar structural rules. In our study, a comparative analysis of the DNA at centromeric regions of Beta species, including cultivated and wild beets, was performed using a set of repetitive DNA sequences. Our results show that these regions in Beta genomes have a complex structure and consist of variable repetitive sequences, including satellite DNA, Ty3-gypsy-like retrotransposons, and microsatellites. Based on their molecular characterization and chromosomal distribution determined by fluorescent in situ hybridization (FISH), centromeric repeated DNA sequences were grouped into three classes. By high-resolution multicolor-FISH on pachytene chromosomes and extended DNA fibers we analyzed the long-range organization of centromeric DNA sequences, leading to a structural model of a centromeric region of the wild beet species Beta procumbens. The chromosomal mutants PRO1 and PAT2 contain a single wild beet minichromosome with centromere activity and provide, together with cloned centromeric DNA sequences, an experimental system toward the molecular isolation of individual plant centromeres. In particular, FISH to extended DNA fibers of the PRO1 minichromosome and pulsed-field gel electrophoresis of large restriction fragments enabled estimations of the array size, interspersion patterns, and higher order organization of these centromere-associated satellite families. Regarding the overall structure, Beta centromeric regions show similarities to their counterparts in the few animal and plant species in which centromeres have been analyzed in detail.