Sabatinos Sarah A, Forsburg Susan L
Department of Molecular and Computational Biology, University of Southern California, 1050 Childs Way, RRI 108, Los Angeles, CA, 90089-2910, USA.
Methods Mol Biol. 2015;1300:79-97. doi: 10.1007/978-1-4939-2596-4_5.
Flow cytometry is an essential tool to monitor DNA content and determine cell cycle distribution. Its utility in fission yeast reflects the ease of sample preparation, the stochiometric binding of the most popular DNA dyes (propidium iodide and Sytox Green), and ability to monitor cell size. However, the study of DNA replication with multicolour flow analysis has lagged behind its use in mammalian cells. We present basic and advanced protocols for analysis of DNA replication in fission yeast by flow cytometry including whole cell, nuclear "ghosts," two-color imaging with BrdU, and estimates of DNA synthesis using EdU.
流式细胞术是监测DNA含量和确定细胞周期分布的重要工具。它在裂殖酵母中的应用体现了样品制备的简便性、最常用DNA染料（碘化丙啶和Sytox Green）的化学计量结合以及监测细胞大小的能力。然而，多色流式分析在DNA复制研究方面的进展落后于其在哺乳动物细胞中的应用。我们提供了通过流式细胞术分析裂殖酵母DNA复制的基础和进阶方案，包括全细胞、核“幽灵”、用BrdU进行双色成像以及使用EdU估计DNA合成。
