Molecular Assays to Investigate Chromatin Changes During DNA Double-Strand Break Repair in Yeast

Abstract

Multiple types of DNA damage, including bulky adducts, DNA single-strand breaks, and DNA double-strand breaks (DSBs), have deleterious effects on the genomes of eukaryotes. DSBs form normally during a variety of biological processes, such as V–D–J recombination and yeast mating type switching, but unprogrammed DSBs are among the most dangerous types of lesion because if left unrepaired they can lead to loss of genetic material or chromosomal rearrangements. The presence of DSBs leads to a DNA damage response involving activation of cell cycle checkpoints, recruitment of repair proteins, and chromatin remodeling. Because many of the proteins that mediate these processes are evolutionarily conserved, the budding yeast, Saccharomyces cerevisiae, has been used as a model organism to investigate the factors involved in the response to DSBs. Recent research on DSB repair has focused on the barrier that chromatin represents to the repair process. In this article, we describe molecular techniques available to analyze chromatin architecture near a defined DSB in budding yeast. These techniques may be of value to experimentalists who are investigating the role of a novel protein in DSB repair specifically in the context of chromatin.