Fig. 3.

Minigene splicing analysis of a novel variant c.550 + 3A > C. The wild-type and mutant genomic fragments of SERPING1 comprising exon 3 and flanking upstream (229 bp) and downstream (255 bp) intron sequences, were cloned into pET01 vector and HepG2 cells were transfected with these minigenes. Capillary electrophoresis of RT-PCR products showed aberrant splicing in nearly 100% of mutated minigene construct transcripts. Several different aberrant transcripts were detected. The most abundant was intron 3 retention followed by exon 3 skipping. Transcripts using cryptic donor splice sites − 10 and + 27 were found in the mutant minigene analysis, whereas they were not detected in the wild type at all. a Minigene analysis results: column A shows transcript proportions resulting from control minigene construct representing c.550 + 3A; column C shows transcript proportions resulting from minigene construct representing c.550 + 3C. b Scheme of the transcripts detected in the minigene analysis. c Scheme of the pET minigene construct. Cryptic splice sites found in the analysis and their exact sequences are displayed. MaxEnt Score values (MES) are specified beneath each splice site