Abstract
The DNA damage response (DDR) is a coordinated cellular response to a variety of insults to the genome. DDR initiates the activation of cell cycle checkpoints preventing the propagation of damaged DNA followed by DNA repair, which are both critical in maintaining genome integrity. Several model systems have been developed to study the mechanisms and complexity of checkpoint function. Here we describe the application of cell-free extracts derived from Xenopus eggs as a model system to investigate signaling from DNA damage, modulation of DNA replication, checkpoint activation, and ultimately DNA repair. We outline the preparation of cell-free extracts, DNA substrates, and their subsequent use in assays aimed at understanding the cellular response to DNA damage. Cell-free extracts derived from the eggs of Xenopus laevis remain a robust and versatile system to decipher the biochemical steps underlying this essential characteristic of all cells, critical for genome stability.