Journal
JOURNAL OF EXPERIMENTAL BOTANY
Volume 67, Issue 21, Pages 6077-6087Publisher
OXFORD UNIV PRESS
DOI: 10.1093/jxb/erw367
Keywords
Arabidopsis; barley; EdU; flow cytometry; maize; replication timing; rice; S-phase duration; wheat
Categories
Funding
- National Science Foundation Plant Genome Research Program (NSF-PGRP) grant [IOS-1025830]
- Direct For Biological Sciences
- Division Of Integrative Organismal Systems [1025830] Funding Source: National Science Foundation
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We estimated S-phase duration for several plant species by following EdU-labeled nuclei from G(1) to G(2) using bivariate flow cytometry. S-phase duration is relatively consistent over a range of genome sizes.The duration of the DNA synthesis stage (S phase) of the cell cycle is fundamental in our understanding of cell cycle kinetics, cell proliferation, and DNA replication timing programs. Most S-phase duration estimates that exist for plants are based on indirect measurements. We present a method for directly estimating S-phase duration by pulse-labeling root tips or actively dividing suspension cells with the halogenated thymidine analog 5-ethynl-2'-deoxyuridine (EdU) and analyzing the time course of replication with bivariate flow cytometry. The transition between G(1) and G(2) DNA contents can be followed by measuring the mean DNA content of EdU-labeled S-phase nuclei as a function of time after the labeling pulse. We applied this technique to intact root tips of maize (Zea mays L.), rice (Oryza sativa L.), barley (Hordeum vulgare L.), and wheat (Triticum aestivum L.), and to actively dividing cell cultures of Arabidopsis (Arabidopsis thaliana (L.) Heynh.) and rice. Estimates of S-phase duration in root tips were remarkably consistent, varying only by similar to 3-fold, although the genome sizes of the species analyzed varied > 40-fold.
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