FIG. 5.

Generation of the DdCenB knockout (KO) cell line and its impact on cytokinesis. (A) Schematic diagram of the creation of the DdCenB knockout construct by PCR and subsequent insertion by homologous recombination. (B) Diagnostic PCR confirming disruption of the DdCenB locus. (C) DdCenB knockout cells show a higher frequency of multinucleation than AX3 and the RFP-DdCenB rescued cell lines. Cells were grown at the described temperatures for 4 days at 200 rpm, fixed with chilled methanol, and stained with DAPI. Nuclei frequencies reflect cell numbers of at least 300 per data point.