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Figure 2


Fig. 2. RNAi of metacaspases in BSF T. brucei. (A). Western blot of cell extracts from an MCA2-MCA3 RNAi clone (left panel) or an MCA5 RNAi clone (right panel), incubated with (+) or without (-) tetracycline (tet) and probed with anti-MCA2-MCA3 or anti-MCA5 antibodies. Asterisk indicates crossreacting 48 kDa protein. (B) Upper panel, growth curve of an MCA2-MCA3 and MCA5 triple RNAi clone with or without induction with tetracycline. Lower panels, western blot of cell extracts from an MCA2-MCA3 and MCA5 triple RNAi clone, with or without induction with tetracycline, probed with anti-MCA2-MCA3 (left) or anti-MCA5 antibodies (right). (C) Upper panel, normal cell-cycle progression in wild-type BSF T. brucei. The kinetoplast division occurs before nuclear division, ultimately followed by cytokinesis. Images illustrate various cell stages with DAPI-stained nuclei (N) and kinetoplasts (K) (white); insets show kinetoplast at higher magnification (blue). Lower panels show nucleus and kinetoplast configurations with or without triple RNAi induction with tetracyline. Others, cells with an abnormal configuration but excluding 2N1K cells. For each time point, 200 cells were analysed. (D) Upper panel, analysis of kinetoplast configuration in 1N1K cells after induction with tetracycline. Bottom panels, aberrant cell-cycle progression in MCA2-MCA3-MCA5 RNAi-induced BSF T. brucei. Images of representative triple RNAi-induced BSF T. brucei. Inset shows kinetoplast at higher magnification. Bars, 10 µm.





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