Single-molecule Analysis of DNA Replication Dynamics in Budding Yeast and Human Cells by DNA Combing

通过 DNA 梳理对芽殖酵母和人类细胞中的 DNA 复制动力学进行单分子分析

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作者:Hélène Tourrière, Julie Saksouk, Armelle Lengronne, Philippe Pasero

Abstract

The DNA combing method allows the analysis of DNA replication at the level of individual DNA molecules stretched along silane-coated glass coverslips. Before DNA extraction, ongoing DNA synthesis is labeled with halogenated analogues of thymidine. Replication tracks are visualized by immunofluorescence using specific antibodies. Unlike biochemical and NGS-based methods, DNA combing provides unique information on cell-to-cell variations in DNA replication profiles, including initiation and elongation. Finally, this assay can be used to monitor the effect of DNA lesions on fork progression, arrest and restart.

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