Chapter title |
Imaging Analysis to Determine Chromatin Binding of the Licensing Factor MCM2-7 in Mammalian Cells.
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Chapter number | 29 |
Book title |
Cell Cycle Control
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Published in |
Methods in molecular biology, January 2014
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DOI | 10.1007/978-1-4939-0888-2_29 |
Pubmed ID | |
Book ISBNs |
978-1-4939-0887-5, 978-1-4939-0888-2
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Authors |
Masayuki Morino, Miyuki Tanaka, Yasushi Shiomi, Hideo Nishitani, Morino, Masayuki, Tanaka, Miyuki, Shiomi, Yasushi, Nishitani, Hideo |
Abstract |
S-CDK and DDK protein kinases initiate DNA replication at replication origins. Prior to the activation of these kinases, origins must become competent for replication by loading MCM2-7 DNA helicase on chromatin. This process is known as replication licensing or pre-replicative complex (pre-RC) formation. After the onset of S phase, however, licensing is inhibited to prevent re-replication of DNA. In this chapter, we describe a method to analyze origin licensing by imaging the chromatin-bound licensing factor MCM2-7. In a normal cell cycle, MCM2-7 is loaded at the end of mitosis or early G1 phase. As S phase progresses, MCM2-7 is dissociated from the replicated regions. When DNA replication is completed, cells in G2 phase have no chromatin-bound MCM2-7. The analysis of chromatin-bound MCM2-7 in each cell provides an insight into cell cycle stage and condition for cell cycle. |
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