Whole-Mount Immunostaining for the Identification of Histone Modifications in the S-Phase Nuclei of Arabidopsis Roots.

Abstract

This chapter describes a method used to analyze the behavior of histone modifications in S phase in Arabidopsis using a whole-mount immunostaining technique. Previous studies have demonstrated that dramatic changes in local chromatin structure are required for the initiation and progression of DNA replication, and that histone modifications play an essential role in the determination of chromatin structure in S phase. Since euchromatic and heterochromatic regions are replicated in distinct S-phase stages, it is important to identify histone modifications at each stage. Here, we introduce a protocol for whole-mount immunostaining combined with 5-ethynyl-2'-deoxyuridine (EdU) staining, which enables the visualization of spatial patterns in histone modifications in the early and late S-phase nuclei of Arabidopsis roots.