Figure 4

Alternative splicing of RVE2 pre-mRNA introduces an in-frame nonsense codon via a PCE event. (A) The schematic representation of the AS event in intron 1 introducing a PCE in the RVE2 transcript. The 5' UTR is shown by a light box, protein coding exons and the PCE are indicated by dark and hatched boxes, respectively. Binding sites for primers F1-R1 and F2-R2 used in RT-PCR are shown by arrows. Normal and premature termination codons are shown by the top and bottom stars. Dashed lines indicate primer portions spanning splice junctions. The gene model is not drawn to scale. (B) The alignment of the 5’ portion of RVE2 cDNA (top) and Sanger sequences of the PCR products (bottom). The PCE sequence is highlighted in grey. The initiation codon is boxed. (C) Quantification of alternatively spliced RVE2 mRNA harboring PCE event using qRT-PCR. The diurnal conditions and the sampling scheme are described in detail in Methods. Arrows indicate the sampling time points (ZT, hours). The relative transcript quantities were calculated using the –ΔΔCt method and CFX Manager software (BioRad). GOG mRNA was used as an internal reference. Vertical bars represent the standard error of the mean.