Cell proliferation, anchorage-independent growth and migration abilities were compromised by the gankyrin knockout. ( A) Growth curves, showing equal numbers of indicated cells (2 × 104) were plated into 12-well tissue culture dishes. Cell numbers were counted at 0, 1, 2, 3, 4, 5 and 6 days. Triplicate wells were used for each time point. The results show the means ± the standard error of the mean for three independent experiments. (B) Cell migration analysis. Cells were seeded onto Oris collagen I-coated plates populated with Oris cell seeding stoppers. After 48 h, the cells were fixed and stained with crystal violet. ( C) Analysis of the levels of cell growth in soft agar. Cells were mixed with soft agar and seeded into 6-well plates; the number of foci was then determined 2~3 weeks later. Data are expressed as the total number of colonies per plate. ( D) Nude mice tumor growth assay. Nude mice (n = 5) were injected subcutaneously in each flank with 5 × 106 cells, and tumor growth was monitored for 3~4 weeks by caliper measurements. Data are shown as the means ± SEM.