FIG. 1.

Disruption of the TLP gene and growth rate of DT40 cells. Disruption of the cTLP gene of DT40 cells by homologous recombination was examined by Northern (A) and Western (B) blotting. The positions of endogenous TLP (cTLP) and ectopically expressed FH-TLP are indicated. +/+, +/−, and −/− represent wild-type, heterozygous TLP mutant, and homozygous TLP mutant (TLP-null) cells, respectively. Three strains of TLP-null cells (strains A, B, and C) were examined. Strain A (lane 3 of panel A and lane 4 of panel B) was obtained by neomycin resistance gene-mediated secondary disruption, and strains B (lane 4 of panel A) and C (lane 5 of panels A and B) were obtained by puromycin resistance gene-mediated secondary disruption. −/−FH-TLP, TLP-null (strain C) cells that stably express FH-TLP. (C) The growth rates of cells of several strains were determined by cell counting at the indicated times. Results for individual strains are shown as average numbers of three different dishes. Dark solid line, wild-type cells. Gray solid lines show two lines of heterozygous TLP-disrupted cells: one for lane 2 of panel A and lane 2 of panel B and the other for lane 3 of panel B. Dotted lines indicate three strains of TLP-null cells. Symbols: ▪, strain A; ×, strain B; •, strain C. (D) Flow cytometry of three different types of DT40 cells. Cell numbers are plotted as a fraction of the DNA contents based on intensities of propidium iodide-dependent fluorescence. −/−, TLP-null (strain C) cells. The percentage of each phase is indicated. The positions of representative G₀/G₁- and G₂/M-phase cells are indicated.