Tamoxifen-induced disruption of NuMA inhibits proliferation of primary embryo fibroblasts. (A) Timeline showing experimental design in which confluent primary cells are treated with 0.1 µM 4-OHT in 2% serum for 48 h. Cells were washed and maintained in 2% serum for 48 h before trypsinization and dilution into media containing 15% serum for subsequent analysis. (B) Conversion of NuMA^flox to NuMA^Δ22 in two independent primary NuMA^flox/flox cell lines carrying the Cre-ER^TM transgene. Recombination was monitored by PCR 48 h after treatment with 4-OHT. (C) qPCR using primers within the floxed region of NuMA was used to measure the efficiency of Cre-mediated recombination in the genomic DNA of fibroblasts. (D) Immunoblotting of NuMA and tubulin in NuMA^flox/flox,Cre-ER^TM and a dilution series of NuMA^+/+,Cre-ER^TM fibroblasts at day 4 of the experimental timeline. (E) Growth curves of primary fibroblasts after 4-OHT–mediated NuMA deletion; n = 3–4 experiments per cell line. Time in days follows timeline shown in A. (F) Mitotic index of primary MEFs treated with 4-OHT. For each genotype, >2,000 cells were counted in two separate cell lines. (G) Duration of mitosis in wild-type (^+/+, Cre) and NuMA-disrupted (^flox/flox, Cre) immortalized embryo fibroblasts. Results represent the mean of two independent experiments. Error bars represent SEM.