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DRB4 over-accumulates after MG132 treatment and in APC/C RNAi lines.

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posted on 24.04.2012 by Katia Marrocco, Marie-Claire Criqui, Jérôme Zervudacki, Gregory Schott, Herfried Eisler, Aude Parnet, Patrice Dunoyer, Pascal Genschik

(A) Characterization of DRB4 overexpressing lines. Northern blot analysis (upper panels) of DRB4 mRNA accumulation in flower extracts from Col-0 and two independent transgenic lines expressing DRB4 under the control of the strong 35S promoter (referred hereafter as lines DRB4 OE-4 and -27). Accumulation of EF1α mRNA is used as a loading control. Western blot analysis (lower panel) performed using an antibody directed against DRB4. The asterisk indicates a non-specific cross-reacting band that can also be used as a loading control. Pictures of 5 week-old Col-0 and DRB4 OE-27 plants. (B) Three week-old DRB4 OE-27 seedlings were incubated in liquid medium supplemented or not with 100 µM MG132. After 4 h and 6 h of incubation, seedlings were collected and total protein extracted. Western blot was performed using antibodies against DRB4 or TSN as a loading control. (C) Western blot analysis of DRB4 accumulation in flower extracts from Col-0, drb4, dcl4, DRB4 OE-27 line, RNAi APC10-38 and RNAi APC6-20 lines. Coomassie staining was used as a loading control (LC).

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