CONCLUSION
In summary, we have carried out a large-scale analysis of exonic variants in SLC12A3 related to GS through bioinformatics predictions and minigene assays. The results indicated that six previously presumed missense variants should be classified as splicing variations, which induced the corresponding exon skipping. These variants either break ESEs and generate new ESSs, or interfere with the canonical splice sites, causing significant pre-mRNA splicing alterations, probably associated with their pathogenicity. This study highlights the important role of assessing the effects of missense variants at the mRNA level in GS, as well as the effectiveness of minigene splicing analysis.