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Journal of Cell Science, Vol 94, Issue 3 471-477, Copyright © 1989 by Company of Biologists
JOURNAL ARTICLES |
AD Mills, JJ Blow, JG White, WB Amos, D Wilcock and RA Laskey
Cancer Research Campaign, Department of Zoology, Cambridge, UK.
Demembranated Xenopus sperm nuclei were induced to replicate synchronously in a low-speed supernatant (LSS) of Xenopus eggs by preincubation in a high-speed supernatant (HSS). DNA replication was observed by incorporation of [alpha-32P]dATP, BrdUTP or biotin-dUTP. Biotin-dUTP incorporation, visualised with fluorescent streptavidin, reveals a striking pattern of replication foci throughout replicating nuclei. We show that this represents a precursor to the bright uniform fluorescence seen later. Confocal microscopic analysis of nuclei fixed early in replication reveals that these foci of DNA replication number about 100-300 for each nucleus and probably represent the replicon clusters already described for tissue culture cells. Foci are evenly distributed throughout the nuclei and are not concentrated at or near the nuclear envelope. Complete replication of each nucleus occurs in an average time of only one hour in this system. Hence we calculate that there must be at least 300-1000 replication forks together in each cluster. Furthermore, pulse labelling at later times in the period of replication reveals a similar pattern of foci indicating that replication forks remain tightly clustered in groups of at least 300 throughout the period of DNA replication.
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D. Rivier and J Rine An origin of DNA replication and a transcription silencer require a common element Science, May 1, 1992; 256(5057): 659 - 663. [Abstract] [PDF]
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R. Laskey, M. Fairman, and J. Blow S phase of the cell cycle Science, November 3, 1989; 246(4930): 609 - 614. [Abstract] [PDF]
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