We have isolated seven DNA fragments from Caenorhabditis elegans that enhance the mitotic segregation of autonomously replicating plasmids in the yeast Saccharomyces cerevisiae. These segregators, designated SEG1-SEG7, behave like isolated yeast chromosomes: they increase the stability and simultaneously lower the copy number of circular plasmids during mitotic growth in yeast. During meiosis, plasmids containing the C. elegans segregators show higher levels of precocious or aberrant disjunction than do plasmids bearing isolated yeast centromeres. Yet one of the segregators improved the meiotic segregation of the parental plasmid. We estimate that there may be as many as 30 segregator sequences in the C. elegans genome, a value that is consistent with the polycentric nature of C. elegans chromosomes. Five of the seven segregators are linked to sequences that are repeated in the worm genome, and four of these five segregators cross-hybridize. Other members of this family of repetitive DNA do not contain segregator function. Segregator sequences may prove useful for probing the structure of centromeres of both C. elegans and S. cerevisiae chromosomes.