Fig. 1. Bud formation is uncoupled from nuclear division in diploid cells treated with IAA. An exponentially growing heterozygous CDC3/CDC3-GFP strain was incubated in YEPD (1% Difco yeast extract, 2% Difco-Bacto Peptone and 2% glucose) plus 0.5% IAA. Samples were removed at 1-hour intervals and stained with DAPI. The samples were examined with DIC microscopy to record overall appearance, and fluorescence microscopy to visualise CDC3-GFP (green) or nuclei (blue). (A) Filaments representative of the changes observed. The time of sampling is indicated in each panel. Arrows in the 1- and 2-hour samples indicate examples of multiple septin patches; the solid arrow in the 8-hour sample (right) indicates a branch forming and the open arrow (left) a double septin ring. Scale bar is 5 µm. The figure does not show examples from all the time points analysed in B and C. (B) The number of buds produced per filament over the 8 hours examined. Over 100 filaments were analysed for each time point. (C) The number of nuclei per filament. Filaments were categorised as indicated, according to whether they had nuclei in a large cell at either end of the filament or a nucleus within the chain of small buds. (D) At the indicated time points, samples were withdrawn and fixed with 2.5% w/v formaldehyde and briefly sonicated. Cell number was determined, counting each clump or filament as a single cell. Filled circles, no IAA; open circles, plus IAA.