An origin-deficient yeast artificial chromosome triggers a cell cycle checkpoint

Checkpoint controls coordinate entry into mitosis with the completion of DN A replication. Depletion of nucleotide precursors by treatment with the dru g hydroxyurea triggers such a checkpoint response. However, it is not clear whether the signal for this hydroxyurea-induced checkpoint pathway is the presence of unreplicated DNA, or rather the persistence of single-stranded or damaged DNA. In a yeast artificial chromosome (YAC) we have engineered a n similar to 170 kb region lacking efficient replication origins that allow s us to explore the specific effects of unreplicated DNA on cell cycle prog ression. Replication of this YAC extends the length of S phase and causes c ells to engage an S/M checkpoint. In the absence of Rad9 the YAC becomes un stable, undergoing deletions within the origin-free region.