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Published 3 July 2006. doi:10.1083/jcb.200604072
The Rockefeller University Press, 0021-9525 $8.00
JCB, Volume 174, Number 1, 89-100
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Article

Isolation of quiescent and nonquiescent cells from yeast stationary-phase cultures



Chris Allen1, Sabrina Büttner4, Anthony D. Aragon1, Jason A. Thomas1, Osorio Meirelles2, Jason E. Jaetao1, Don Benn1, Stephanie W. Ruby3, Marten Veenhuis5, Frank Madeo4, and Margaret Werner-Washburne1

1 Department of Biology, 2 Department of Mathematics and Statistics, and 3 Department of Molecular Genetics and Microbiology, Health Sciences Center, University of New Mexico, Albuquerque, NM 87131
4 Institute of Molecular Biology, Biochemistry, and Microbiology, Karl-Franzens University, 8010 Graz, Austria
5 Department of Eukaryotic Microbiology, University of Groningen, Kerklaan 30, 9750 AA Haren, Netherlands

Correspondence to Margaret Werner-Washburne: maggieww{at}unm.edu

Quiescence is the most common and, arguably, most poorly understood cell cycle state. This is in part because pure populations of quiescent cells are typically difficult to isolate. We report the isolation and characterization of quiescent and nonquiescent cells from stationary-phase (SP) yeast cultures by density-gradient centrifugation. Quiescent cells are dense, unbudded daughter cells formed after glucose exhaustion. They synchronously reenter the mitotic cell cycle, suggesting that they are in a G0 state. Nonquiescent cells are less dense, heterogeneous, and composed of replicatively older, asynchronous cells that rapidly lose the ability to reproduce. Microscopic and flow cytometric analysis revealed that nonquiescent cells accumulate more reactive oxygen species than quiescent cells, and over 21 d, about half exhibit signs of apoptosis and necrosis. The ability to isolate both quiescent and nonquiescent yeast cells from SP cultures provides a novel, tractable experimental system for studies of quiescence, chronological and replicative aging, apoptosis, and the cell cycle.

Abbreviations used in this paper: AnnV, Annexin V; ANOVA, analysis of variance; DHE, dihydroethidium; FUN-1, 2-chloro-4-(2,3-dihydro-3-methyl-[benzo-1, 3-thiazol-2-yl]-methylidene)-1-phenylquinolinium iodide; PI, propidium iodide; ROS, reactive oxygen species; SP, stationary phase.


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