The U2 snRNP participates in the splicing of the Tsta3 minor intron in the presence of functional U12 snRNPs. (A) Representative gel of RT-PCR experiments analyzing simultaneously the amount of unspliced and spliced products obtained with the different reporters. Reporter names and size (in base pairs) of the amplified products are indicated on the left and the schematic structure of the splicing products is shown on the right. Preincubation of the splicing extracts with LNAs targeting the U2 (U2, 2 µM), U12 (U12, 2 nM), or U12 + U2 snRNAs is indicated. The faint band detected immediately below the Slc12a4 and Vac14 unspliced products likely corresponds to a spliced–unspliced heteroduplex RNA. (B) Splicing efficiency (in %) determined for the different reporter substrates following incubation in control (Ctl) or LNA-pretreated extracts. (C) Representative gel of RT-PCR experiments analyzing simultaneously the amount of unspliced and spliced products obtained with the Slc12a4 and Tsta3 reporters. Preincubation of the splicing extracts with LNAs targeting the U2 (U2, 2 µM) or U12 (U12, 2 µM) snRNAs is indicated. (D) Splicing efficiency (in %) determined for the different reporter substrates following incubation in control (Ctl) or LNA-pretreated extracts. Statistical significance was calculated using the Student t-test (n = 3; (*) P < 0.05; (**) P < 0.01; (***) P < 0.001; (****) P < 0.0001).
