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Methods. 2007 Feb;41(2):222-31.

Analyzing the ATR-mediated checkpoint using Xenopus egg extracts.

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  • 1Department of Molecular Pharmacology, Stanford University, 318 Campus Drive , Stanford, CA 94305-5441, USA.

Abstract

Our knowledge of cell cycle events such as DNA replication and mitosis has been advanced significantly through the use of Xenopus egg extracts as a model system. More recently, Xenopus extracts have been used to investigate the cellular mechanisms that ensure accurate and complete duplication of the genome, processes otherwise known as the DNA damage and replication checkpoints. Here we describe several Xenopus extract methods that have advanced the study of the ATR-mediated checkpoints. These include a protocol for the preparation of nucleoplasmic extract (NPE), which is a soluble extract system useful for studying nuclear events such as DNA replication and checkpoints. In addition, we describe several key assays for studying checkpoint activation as well as methods for using small DNA structures to activate ATR.

PMID:
17189864
PMCID:
PMC2657337
DOI:
10.1016/j.ymeth.2006.07.024
[PubMed - indexed for MEDLINE]
Free PMC Article
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