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. 2017 Aug 10;45(18):10481–10491. doi: 10.1093/nar/gkx676

Figure 3.

Figure 3.

M1BP is required for RP gene transcription in vitro. (A) M1BP-probed (top) and Rpb3-probed (bottom) western blot of purified His-M1BP and undepleted, mock-depleted or M1BP-depleted nuclear extracts from 0–12 h embryos. A total of 10 or 30 ng purified His-M1BP and 20 or 60 µg of each extract type was loaded for SDS-PAGE western blot analysis. (B and C) Primer extension analysis of transcripts produced from the (B) RpL30 or (C) RpLP1 promoter in embryo nuclear extracts. The bracketed region denotes the same TSS region described in Figure 2A. Each transcription reaction was performed in duplicate. Lanes 1 and 2: mock-depleted extract supplemented with dialysis buffer. Lanes 3 and 4: mock depleted extract supplemented with recombinant M1BP. Lanes 5 and 6: M1BP-depleted extract supplemented with dialysis buffer. Lanes 7 and 8: M1BP-depleted extract supplemented with enough recombinant M1BP to replace the amount that was immunodepleted. (D) Coomassie-stained SDS-PAGE analysis of purified, N-terminally His-tagged M1BP expressed in and purified from Escherichia coli.