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Updated: Aug 12, 2026

An Efficient Method for Quantitative, Single-cell Analysis of Chromatin Modification and Nuclear Architecture in Whole-mount Ovules in Arabidopsis
Published on: June 19, 2014
A standardized method for the three-way differential staining of plant chromosomes and the scoring of SCEs per cell
P Escalza1, J Piñero, F Cortés
1Departamento de Biología Celular, Facultad de Biología, Sevilla, Spain.
Abstract:
We have made use of 2 alternative methodologies to obtain 3-way differential staining (TWD) in third-mitosis (M3) chromosomes of Allium cepa, which involve different uptakes of bromodeoxyuridine (BrdU) through 3 cell divisions, in order to evaluate the sister-chromatid exchanges (SCE) frequency on a per-cell-cycle basis. The main innovation introduced by us to the original protocols previously reported has been the use of 5-fluorodeoxyuridine (FdU) to inhibit endogenous thymidine (dT) synthesis. By using different [BrdU]:[dT] ratios in the presence of FdU the relative incorporation of BrdU into replicating DNA can be controlled. The use of 2 different approaches to obtain 3-way differentiation of sister chromatids allowed us to evaluate the role of BrdU in the induction of SCEs in our system. Both procedures rendered nearly 100% of M3 chromosomes showing TWD. An additional advantage of our methodologies is their high degree of reproducibility.
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