Fig. 4. Experimental procedure and selection of daughter cells for imaging. (a) Scheme depicting the experimental procedure. Cells were microinjected during the first S phase with a mixture of the GFP-PCNA expression plasmid and Cy3-dUTP. After 1.5 hours, some cells were microinjected again with Cy5-dUTP. Cells were imaged during the next S phase after cell division. (b) Schematic drawing of nuclei selected for imaging. During the imaging period following cell division, labelled cells were at different cell cycle and S-phase stages. Cells not in S phase at all (left, uniform distribution of GFP-PCNA) or not at that stage of S phase that corresponded to the DNA-labelling pattern (middle, GFP-PCNA localizing in different nuclear regions to the labelled DNA) were not chosen for imaging. Only those cells where GFP-PCNA foci and labelled DNA foci occupied similar nuclear regions were imaged (right).