Fig. 3

The CENP-A domain occupies a similar proportion of alpha satellite DNA throughout the cell cycle. Chromatin fibers were isolated from three male cell lines that were synchronized in G1, G1/S, and S phase. Fibers were immunostained for CENP-A, followed by hybridization with alpha satellite probes to the centromeres of HSAX (DXZ1) or HSAY (DYZ3). Each alpha satellite array varied in length, as previously determined by molecular analyses [18]. a, b Results of chromatin fiber CENP-A immunostaining (green) and alpha satellite FISH (red). Scale bars are 5 μm. c, d Percentage of alpha satellite occupied by CENP-A chromatin throughout the cell cycle. The lengths of the FISH signal and CENP-A signal were measured, and the proportion of the alpha satellite array occupied by CENP-A chromatin was calculated for each cell cycle stage. PF primary fibroblast, CF cancer fibroblast, LCL lymphoblastoid cell line