Table 1.

Studies that explored alternative splicing by using pDESTsplice or pSpliceExpress.

Reference	Vector	Issue	Context	Study Topic	ASE	Special Features
Abdulhay et al., 2019 [55]	pSpliceExpress	Disease-related	Hematopoiesis	Genetic variants	Intron retention	Investigations on an intronic mutation disrupting the activity of snRNP U2
Alaa el Din et al., 2015 [46]	pSpliceExpress	Disease-related	Hereditary hemorrhagic telangiectasia syndrome	Genetic variants	Alt. splice site, Intron retention	Examination of the pathogenicity of genetic variants and their influence on splicing
Bartosovic et al., 2017 [49]	pDESTsplice	Physiological role	RNA modification via FTO demethylase	RNA modification	Exon skipping	Comparison of splicing regulation in FTO knock-out cells vs. wildtype cells
Beaman et al., 2019 [57]	derivate of pSpliceExpress	Disease-related	Urinary bladder disease	Genetic variants	N/A	Case report
Cao et al., 2020 [62]	pSpliceExpress	Physiological role	pMEIs	Genetic variants	Exon skipping	Experimental validation of pMEI sQTLs based on data from the GTEx project
Carvill et al., 2018 [50]	pDESTsplice	Disease-related	Dravet syndrome and related genetic epilepsies	Genetic variants	Intron retention	Analysis of a new genetic variant identified by genome sequencing in a patient
Chase et al., 2020 [60]	pSpliceExpress	Disease-related	Myeloid neoplasms	Genetic variants	Exon skipping	Effect of mutations on methylation activity and the splicing process
Dupont et al., 2019 [58]	pSpliceExpress	Disease-related	Diseases linked to cilium	Genetic variants	Exon skipping	Comparison of IFT52 mutations in fetuses with distinct phenotypes
Ellingford et al., 2019 [59]	derivate of pSpliceExpress	Disease-related	Rare monogenic disorders	Genetic variants	Alt. splice site, Cryptic splice site, Exon skipping, Intron retention	Experimental set-up to examine accuracy of in silico variant prioritization strategies
Kishore et al., 2010 [41]	pSpliceExpress	Disease-related	Prader–Willi syndrome	Splicing regulators	Exon skipping, Intron retention	Cotransfection of target minigenes and SNORD 115 expression constructs
Knapp et al., 2020 [63]	pSpliceExpress	Disease-related	Meier–Gorlin syndrome	Genetic variants	Intron retention	Identification of novel genetic variants in genes that cause disease
Legendre et al., 2018 [51]	pSpliceExpress	Disease-related	CHARGE syndrome	Genetic variants	Intron retention	Branch point analyses
Listerman et al., 2013 [44]	pSpliceExpress	Disease-related	Cancer biology	Splicing regulators	Exon skipping	SRSF11, hnRNPH2 and hnRNPL regulate TERT exon 7/8 skipping
Mattison et al., 2018 [53]	pDESTsplice	Disease-related	Epilepsy	Genetic variants	Exon skipping	Splicing studies on genetic variants discovered in patients
Mutai et al., 2020 [64]	pSpliceExpress	Disease-related	Hereditary hearing loss	Genetic variants	Exon skipping	Combination of minigene assays and functional analyses in cochlear tissues
Payer et al., 2019 [54]	pSpliceExpress	Physiological role	Alu polymorphisms	Genetic variants	Exon skipping	Influence of Alu element polymorphisms on splicing
Rittore et al., 2014 [45]	pSpliceExpress	Disease-related	Inflammatory diseases	Genetic variants	Exon skipping	Assessment of combinatorial effects of SNPs
Reference	Vector	Issue	Context	Study Topic	ASE	Special Features
Scott et al., 2012 [43]	pDESTsplice	Disease-related	Cystic fibrosis	Genetic variants	Cryptic splice site, Exon skipping	Selection of genetic variants for experimental testing via bioinformatic tools
Starokadomskyy et al., 2016 [48]	pSpliceExpress	Disease-related	X-linked late pigmentary disorder	Genetic variants	Intron retention	Investigations on an intronic mutation causing a rare X-chromosomal disease
Sumanasekera et al., 2012 [42]	pSpliceExpress	Disease-related	Ceramide-mediated splicing, Cancer drug	Splicing regulators	Alt. splice site, Exon skipping	Influence of C6 pyridinium ceramide on splicing
Tang et al., 2020 [65]	pDESTsplice	Disease-related	Alzheimer’s disease	Genetic variants	N/A	Investigations of genotype-dependent splicing efficiencies
Thomas et al., 2020 [61]	pSpliceExpress	Disease-related	Mandibulofacial dysostosis Guion–Almeida type	Genetic variants	Cryptic splice site, Exon skipping, Intron retention	Investigations on pathogenic variants altering splicing of the human EFTUD2 gene and the yeast homolog SNU114
Varga et al., 2019 [56]	pSpliceExpress	Disease-related	Autosomal dominant sensorineural hearing loss	Genetic variants	Exon skipping	Case report
Wang et al., 2018 [52]	pSpliceExpress	Physiological role	Mammalian cerebellar development	RNA modification	Exon skipping	Aberrant splicing due to METTL3-mediated m6A modification
Xiao et al., 2016 [47]	pSpliceExpress	Physiological role	Splicing regulatory factors, RNA-binding proteins	Splicing regulators	Exon skipping	Splicing regulation of ZNF638 upon knockdown of YTHDC1, SRSF3 or SRSF10

We compiled 25 articles (sorted by author name) that described a minigene splicing assay on the basis of the pDESTsplice or pSpliceExpress vector systems. The physiological role of splicing variants or their relation to diseases was investigated. The influences on splicing by genetic variants, RNA modifications or splicing regulatory factors were examined in the 25 studies. The investigated alternative splicing patterns can be distinguished into 4 different types of ASE: alternative splice sites, cryptic splice sites, exon skipping, and intron retention. The specific research context and special features of the individual studies were also recorded in the table. Alt.: alternative, ASE: alternative splicing events, N/A: not available, pMEIs: polymorphic mobile element insertions, SNP: single-nucleotide polymorphism, sQTLs: splicing quantitative trait loci.