The SeqSplice workflow. (A) Experimental design of the minigene splicing assays. Synthetic BRCA1 and BRCA2 exons (pink) and flanking intronic sequences (gray) are cloned in between the pSPL3 vector exons, and variants are introduced via mutagenesis. WT and variant minigene constructs are transfected into HS578T and MDA-MB-231 cell lines, followed by RNA sequencing of the produced transcripts. Transcript proportions are calculated using our custom SeqSplice software package. (B) A conceptual breakdown of the steps involved in the SeqSplice RNA isoform identification and quantitation.