Figure 4

Microarchitecture of ESS activity in exon 26 sequences derived from fragment 6. (A) In vitro splicing assay of DNA ligase III C-ß splicing reporter, tagged with 25-mers derived from fragment 6 (1 hour splicing at 30°C). 10% PAGE shown. Lanes Cß = DNA ligase III C-ß construct without ESS; Cß ex27 1–25 = Cß + nt 1–25 of exon 27; Cß REP = Cß + native ESS; Cß IGF-I ex3 1–25 = Cß + nt 1–25 of exon 3 from IGF-I; DL3 Cß ex 26 lanes = nts of exon 26 tested are shown above each lane; lane 2319–2343 encompasses the RNA editing site and mooring sequence (ED); ED sequence mutations T9A, T9C, A11C, G15C, A19T shown in right-hand five lanes. Adjacent cartoons show identity of bands. (B) Quantification of splicing efficiency, normalised to DNA ligase III C-ß reporter. Bars show splicing efficiency from n=4 experiments with DNA ligase III C-ß = 100%. Error bars show S.E.M. Overall one-way ANOVA shows p <0.0001. Comparison to DNA ligase III C-ß for each construct by Bonferroni’s multiple comparison test: statistically significant differences from efficiency of DNA ligase III C-ß splicing shown by *= p <0.01, **= p <0.001.