We identified a novel causative mutation of NTG in gene X by performing WES. Moreover, this novel mutation exhibits the gain of aberrant mRNA splicing that leads to retention of intros or skipping exon and eliminates full-length protein using several different cell culture models, including patient-derived iPSCs. The aberrantly spliced variants affected protein production and altered subcellular localization. Furthermore, we developed and characterized the knock-in and knockout mouse models. Both knock-in and knockout mice had decreased RGCs and retinal thickness. Last, we differentiated iPSCs into RGCs by inhibiting SMAD and Wnt pathways. The mutational effects of the novel gene were analyzed in purified iPSC-RGCs.