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. 2013 Jun 6;50(5):661–674. doi: 10.1016/j.molcel.2013.05.011

Figure 1.

Figure 1

Replication Factories Are Initially Formed at Centromeric Regions

(A) Replication factories are initially formed near the SPB. GFP-PCNA (T8375 [i]) and POL1-GFP (T7300 [ii]) cells with SPC42-mCherry NIC96-mCherry (visualizing SPBs and nuclear pores [NPs]) were observed in asynchronous culture. T7300 cells also had NET1-mCherry (nucleolus marker). We set 0 s arbitrarily. The scale bar represents 1 μm.

(B) Replication factories are initially formed, overlapping with the kinetochore cluster. Cells (T10459) with GFP-PCNA SPC42-mCherry NIC96-mCherry NDC80-CFP (visualizing kinetochores [KTs]) were observed in asynchronous culture. White lines represent the nucleus and the SPB. CFP, cyan fluorescent protein.

(C) Replication factories are initially formed on centromeres when they are detached from a spindle pole. Cells (T8819) with GFP-PCNA MTW1-CFP NDC80-CFP (KTs-CFP) SPC42-mCherry NIC96-mCherry mad2Δ bub2Δ were arrested with α factor and released into medium with nocodazole. Kinetochores were frequently detached from a spindle pole after going through S phase in the presence of nocodazole. mad2Δ bub2Δ allowed cells to exit from mitosis in the absence of microtubules. Cells were observed after cytokinesis (125 min after release from α factor).