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Journal of Cell Science, Vol 107, Issue 11 3045-3053, Copyright © 1994 by Company of Biologists

JOURNAL ARTICLES

Initiation of DNA synthesis by nuclei from scrape-ruptured quiescent mammalian cells in high-speed supernatants of Xenopus egg extracts

M Hola, S Castleden, M Howard and RF Brooks
Division of Biomedical Sciences, King's College, Randall Institute, London, UK.

Demembranated sperm heads, detergent-isolated somatic nuclei and even naked DNA are efficiently replicated in cytoplasmic extracts of activated amphibian eggs, but only after nuclear assembly and the formation of an intact nuclear envelope. DNA synthesis has not previously been shown to be initiated in high-speed (200,000 g) supernatants of egg cytoplasm because they are depleted of the vesicular material required to support nuclear envelope formation. Here we show that mammalian nuclei prepared by scrape-rupture are able to initiate DNA replication in such high-speed supernatants. These nuclei begin DNA synthesis asynchronously. This asynchrony cannot be attributed to differences in the time taken for nuclear assembly. Instead, we suggest that the asynchrony reflects intrinsic differences between nuclei and that these differences are a major cause of cell cycle variability. Our demonstration of initiation in high-speed supernatants now enables the initiation of eukaryotic DNA synthesis to be studied independently of nuclear assembly.


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W Sun, M Hola, K Pedley, S Tada, J. Blow, I. Todorov, S. Kearsey, and R. Brooks
The replication capacity of intact mammalian nuclei in Xenopus egg extracts declines with quiescence, but the residual DNA synthesis is independent of Xenopus MCM proteins
J. Cell Sci., January 2, 2000; 113(4): 683 - 695.
[Abstract] [PDF]


© The Company of Biologists Ltd 1994