목적: OTOA encodes Otoancorin, required for the development of the tectorial membrane in the inner ear. Alterations in the gene cause non-syndromic sensorineural hearing loss (SNHL)(DFNB22). Previous study revealed that disrupted glycosylphosphatidylinositol (GPI) anchorage of Otoancorin played an important role in the pathogenesis of DFNA22. A 10-month-old boy visited the clinic presenting with bilateral moderate SNHL. Routine short-read sequencing (SRS) resulted in two potential candidate variants of OTOA: p.Gln589Argfs*55 (known pathogenic variant) and p.Glu787*. p.Glu787* is expected to disrupt the GPI anchorage, which should be pathogenic according to the previous report if it is real. However, too high minor allele frequency (0.23) in Korean population was highly against being considered as pathogenic. To address the issue that p.Glu787* might reside in pseudogene (OTOAP1) not in OTOA, thus not exerting pathogenic potential in reality. We performed the long-read sequencing (LRS) and further generated the Knock-in (KI) mouse of p.Glu787* to directly prove its pathogenicity. 방법:DNA samples of proband and parents were collected and SRS & LRS were performed to identify the causative variants. To confirm whether p.Glu787* resides in OTOAP1 or not, data from SRS and LRS were compared using IGV, BLAST and other bioinformatics tools. KI mouse (OtoaE787*E787*) was generated. ABR was performed to assess the hearing and H&E, IHC & electron microscopy (EM) were done to observe the immunohistology in the cochlea. 결과:LRS and bioinformatics analysis confirmed the presence of p.Glu787* in the OTOAP1. ABR at P28 showed severe hearing loss and histologic study revealed detached tectorial membrane. 결론:LRS revealed the pseudogene contamination that made genetic diagnosis difficult. KI mouse study confirmed the pathogenic potential of p.Glu787* if it is present in the true Otoa. Attention should be taken to interpret the NGS data in case of genes with possible pseudogene contamination.