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J. Cell Biol.,
Volume 144, Number 5, March 8, 1999 813-822
Sir William Dunn School of Pathology, University of Oxford, Oxford, OX1 3RE, United Kingdom
Individual chromosomes are not directly visible within the interphase nuclei of most somatic cells;
they can only be seen during mitosis. We have developed a method that allows DNA strands to be observed
directly in living cells, and we use it to analyze how mitotic chromosomes form. A fluorescent analogue (e.g.,
Cy5-dUTP) of the natural precursor, thymidine triphosphate, is introduced into cells, which are then grown on
the heated stage of a confocal microscope. The analogue is incorporated by the endogenous enzymes into
DNA. As the mechanisms for recognizing and removing the unusual residues do not prevent subsequent
progress around the cell cycle, the now fluorescent
DNA strands can be followed as they assemble into
chromosomes, and segregate to daughters and granddaughters. Movies of such strands in living cells suggest
that chromosome axes follow simple recognizable paths
through their territories during G2 phase, and that late
replicating regions maintain their relative positions as
prophase chromosomes form. Quantitative analysis
confirms that individual regions move little during this stage of chromosome condensation. As a result, the
gross structure of an interphase chromosome territory
is directly related to that of the prophase chromosome.
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