Splicing complementation of ΔMS extracts. (A) Splicing was performed at 25°C for 30 min in ΔMS extracts without (lane 1) or with the addition of various amounts of ΔM (lanes 2–5) or ΔMS (lanes 6–9) extracts. Lanes 2 and 6: 0.13 µg/µL; lanes 3 and 7: 0.4 µg/µL; lanes 4 and 8: 1.3 µg/µL; lanes 5 and 9: 4 µg/µL. (Compl Ext) Complementing extract; ΔM, MUD2 deletion; ΔMS, MUD2, SUB2-double deletion. (B) The exon ligation efficiency of reactions from (A), calculated as the molar ratio of mRNA to the total of mRNA plus pre-mRNA. Data represent mean values from three independent experiments with standard deviation (SD) indicated. (C) Splicing was performed in ΔMS extract without (None) or with the addition of 2 μg/μL ΔM (ΔM) or ΔMS (ΔMS) extracts, together with various amounts of M2M5 proteins. Data represent mean values from three independent experiments with standard deviation (SD) indicated. M2M5, recombinant Mud2-Msl5 dimer; ΔM, MUD2 deletion; ΔMS, MUD2, SUB2-double deletion.
