Figure 4 – rAAV8
producing cells treated with nocodazole arrest in G2/M phase. Flow
cytometry histograms triggered against propidium iodide signal for cells
show the relative abundance of cells in either G1 (blue), S (orange) or
G2/M (green) phases of the cell cycle in the absence (A ) or
presence (B ) of 4 µM nocodazole addition at 0 HPT. Ratio of
cells in G2/M phase to G1 phase at 72 HPT with 4 µM nocodazole added at
0, 4, 24, 48 HPT or untreated (Ctrl) (C ). Representative
fluorescent composite images of rAAV-producing cells fixed 24 HPT show
staining of the fibrillar component of the nucleolus with
anti-fibrillarin (red) and nuclear DNA with DAPI (blue). Cells
displaying disorganised nucleolar morphology are indicated by white
arrows in non-treated cells (D ) and cells treated 4 hours
post-transfection with 4 µM nocodazole (E ). Images taken using
100 x oil objective. Scale bar 10 µm. Data shown in (C ) are
mean ± SEM for three biological replicates.