Splice site strength and order influence 3′ alternative cassette selection. (A) An overview of the various mutant constructs generated to assess the effects of splice site strength on 3′ alternative cassette selection. Red arrow depicts increasing or decreasing strength of the splice site. The WT and modified 3′ splice site sequences are presented with the scores shown on the right. Uppercase letters indicate exon sequences; lowercase letters indicate intron sequences. Mutated nucleotides are marked in red. (B) Effects of splice site strength mutations on the inclusion of alternative cassette LCx or LCy. RT-PCR was performed to detect the RNA splicing pattern. (C) Effects of the mutations on the inclusion of alternative cassette LCx or LCy. (D) Schematic diagrams of the mutant constructs used to assess the effects of alternative cassette order on its selection. (E) Effects of alternative cassette order on its selection. LCx and LCy inclusion was switched when their order was exchanged. (F) The frequency of alternative 3′ cassette is dependent on its position (order). Data are expressed as the mean ± SD from three independent experiments.
