Multiple approaches to study S. cerevisiae Rad9, a prototypical checkpoint protein

Aisling M O'Shaughnessy; Muriel Grenon; Chris Gilbert; Geraldine W-L Toh; Catherine M Green; Noel F Lowndes

doi:10.1016/S0076-6879(05)09008-7

Multiple approaches to study S. cerevisiae Rad9, a prototypical checkpoint protein

Methods Enzymol. 2006:409:131-50. doi: 10.1016/S0076-6879(05)09008-7.

Authors

Aisling M O'Shaughnessy¹, Muriel Grenon, Chris Gilbert, Geraldine W-L Toh, Catherine M Green, Noel F Lowndes

Affiliation

¹ Department of Biochemistry, National University of Ireland, Galway, Ireland.

PMID: 16793399
DOI: 10.1016/S0076-6879(05)09008-7

Abstract

The Saccharomyces cerevisiae RAD9 checkpoint gene is the prototypical checkpoint gene and is required for efficient checkpoint regulation in late G1, S, and at the G2/M cell cycle transition following DNA damage. Rad9 is required for the activation of Rad53 after damage and has been proposed to have roles in lesion recognition as well as DNA repair and the maintenance of genome stability. Here we describe methodology suitable for the study of G1, intra-S, and G2/M checkpoints in budding yeast, the analysis of Rad9/Rad53 phospho-forms, the biochemical analysis of Rad9 and Rad53, the fractionation of soluble and chromatin associated proteins, including Rad9, and the live cell imaging of GFP tagged Rad9.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Blotting, Western
Cell Cycle / genetics*
Microscopy, Fluorescence
Saccharomyces cerevisiae / genetics*
Saccharomyces cerevisiae Proteins / genetics*

Substances

Saccharomyces cerevisiae Proteins