Figure EV1. The effect of combined nocodazole and cytochalasin‐B treatment on the distribution of endogenously labelled PCM1‐GFP in DT40 cells.

1. Diagram showing the GFP construct used to target the chicken PCM1 locus at the C‐terminus on both alleles, by homologous recombination. Highlighted the Sal1 and BamH1 sites used for restriction digestion to clone the LA (Left Arm) and the RA (Right Arm) and to replace the resistance cassette. Clones were screened for antibiotic resistance genes blasticidin (Blasti), puromycin (Puro) or Histidinol (His). LoxP sites flanking the resistance cassette are represented by red triangles. The dashed lines indicate the sites of recombination and integration in the PCM1 locus. Confirmation of targeting was carried out by Western blotting, as shown in Fig 1B–D.
2. Representative immunofluorescence images of cell lines with genotypes as indicated, treated with both nocodazole (2 μg/ml) and cytochalasin‐B (1 μg/ml). DMSO‐treated cells were used as a control (DMSO, upper panels). Treatments were carried out for 2 h, and cells were co‐stained with antibodies against GFP (green) and γ‐tubulin (red). DNA is in blue. Images correspond to maximum intensity projections of confocal micrographs. Asterisks mark cells with dispersed satellites. Note that drug treatment leads to an increase in large and a decrease in small satellite granules in all three genotypes, but the effects are more prominent in acentriolar than in WT cells. Scale bars: 5 μm.