Figure 4

The effect of hSlu7 on tandem splice site selection. HeLa cells were transfected with the RNAi oligonucleotide directed against either hSlu7 or luciferase (control). (A) After 48 h, total protein was extracted, separated by 10% SDS-PAGE, transferred to a membrane and probed for hSlu7 and actin. (B) The total RNA was collected, followed by RT-PCR using specific primers. The alternative splicing of the DDO-1 gene was affected by the hSlu7 protein concentration (positive control). GAPDH, a housekeeping gene, was amplified in each sample to confirm that approximately the same amount of cDNA was used for each reaction. (C) and (D) Analysis of 3' wobble splicing of three endogenous genes (FOXM1: NM_021953, PGAM5: NM_138575 and METTL9: NM_016025) and four minigenes (RAGE: NM_014226, SIPA1L1: NM_015556, ARID1A: NM_018450 and AG(T)₉CAG) in hSlu7 knockdown HeLa cells using capillary electrophoresis (upper panel). The relative percentage of the two isoforms was calculated using GeneScan 3.7 (lower panel).