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Analyzing DNA Replication Checkpoint in Budding Yeast

  • Nicole Hustedt
  • Kenji Shimada
Part of the Methods in Molecular Biology book series (MIMB, volume 1170)

Abstract

Checkpoints are conserved mechanisms that prevent progression into the next phase of the cell cycle when cells are unable to accomplish the previous event properly. Cells also possess a surveillance mechanism called the DNA replication checkpoint, which consists of a conserved kinase cascade that is provoked by insults that block or slow down replication fork progression. In the budding yeast Saccharomyces cerevisiae, the DNA replication checkpoint controls the timing of S-phase events such as origin firing and spindle elongation. This checkpoint also upregulates dNTP pools and maintains the replication fork structure in order to resume DNA replication after replication block. Many replication checkpoint factors have been found to be tumor suppressors, highlighting the importance of this checkpoint pathway in human health. Here we describe a series of protocols to analyze the DNA replication checkpoint in S. cerevisiae.

Key words

DNA replication checkpoint Mec1 Rad53 Kinase assay Budding yeast 

Notes

Acknowledgements

We thank Dr. Susan M. Gasser for critical reading and helpful suggestions. Support was gratefully received from the Novartis Research Foundation, the Swiss Cancer League, and the FP7 Marie Curie Network, Imaging the DNA Damage Response (Image DDR).

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Friedrich Miescher Institute for Biomedical ResearchBaselSwitzerland

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