Fig. 5: Minigene-splicing assay.
From: Elucidation of the aberrant 3′ splice site selection by cancer-associated mutations on the U2AF1
a Schematic diagram of ATR minigene designed for the splicing assay, which has different nucleotide A or T at the −3 position of the 3′ splice site. The minigene exons are shown as boxes, and introns as solid lines. Two different pre-mRNA splicing patterns are shown as exon inclusion and exon skipping. The RNA product of the exon inclusion is derived from the removal of two introns and junction of three exons, whereas that of the exon skipping is produced by the junction of exons at both ends. b Gel electrophoresis of RT-PCR products from HEK293 cells expressing WT or mutants of U2AF1. Upper fragments correspond to the product by the exon inclusion, and lower fragments correspond to that by the exon skipping. Raw image data of gel electrophoresis are shown in Supplementary Fig. 13. c The ratio of ATR exon inclusion and the skipping is quantified from the result of (b), and compared based on the amount from cells expressing WT with TAG minigene.
