We have examined the effect of physical length on the mitotic segregation of artificial chromosomes and fragments of natural yeast chromosomes. Increasing the length of artificial chromosomes decreases the rate at which they are lost during mitosis. We have made fragments of chromosome III by integrating new telomeres at different positions along the length of the chromosome. Chromosome fragments of 42 and 72 kb behave like artificial chromosomes: they are lost in mitosis much more frequently than natural chromosomes. In contrast, a chromosome fragment of 150 kb is as mitotically stable as the full-length chromosome from which it is derived. The structural instability of a short dicentric artificial chromosome demonstrates that, although short artificial chromosomes segregate poorly in mitosis, they do attach to the mitotic spindle. We discuss these results in the context of a model in which chromosome segregation is directed by the intercatenation of the segregating DNA molecules.