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. 2017 Feb 9;8(16):26027–26040. doi: 10.18632/oncotarget.15215

Figure 3. Functional analysis of Boff-p210-edited cells.

Figure 3

(A) Western blot analysis of BCR-ABL expression. Abl expression (140 kDa) was observed in all cells. A 210 kDa band, corresponding BCR-ABL expression was observed in unedited Boff-p210 cells (parental and sgRNA mock controls) and in Bcr-Abl-EP cells. In contrast, Bcr-Abl-SC and Baf/3 cells showed no expression of BCR-ABL. (B) Annexin V/propidium iodide labeling of Boff-p210 cells after four days in culture in the presence or absence of IL-3. Unedited and mock sgRNA-expressing cells showed IL-3-independent growth. Baf/3 as Bcr-Abl-EP cells showed greater annexin V labeling after IL-3 withdrawal. This effect was stronger in cell lines derived from a single-edited cell. (C) Flow cytometric analysis of Bcr-Abl-EP cells. No changes were observed in the cell cycle phases of Boff-p210 or the mock cells, grown with or without IL-3. High frequencies of cells in the subG0 phase were found in the single edit derived-cells cultured without IL-3, as in Baf/3 cells. (D) Quantification of annexin V labeling (Left graph, represented as mean ± SEM; ***p < 0.001) and percentage of Boff-p210-positive cells (right graph).