Objective: The objective of this study was to pinpoint pathogenic genes and assess the mutagenic pathogenicity in two pediatric patients with hereditary spherocytosis.Methods: We utilized whole-exome sequencing (WES) for individual analysis (case 1) and family-based trio analysis (case 2). The significance of the intronic mutation was validated through a Minigene splicing assay and supported by subsequent in vitro experiments.Results: Both probands received a diagnosis of hereditary spherocytosis. WES identified a novel ANK1 c.1504-9G>A mutation in both patients, causing the retention of seven nucleotides at the 5′ end of intron 13, as substantiated by the Minigene assay. This variant results in a premature stop codon and the production of a truncated protein. In vitro studies indicated a reduced expression of the ANK1 gene.Conclusion: The novel ANK1 c.1504-9G>A variant is established as the causative factor for hereditary spherocytosis, with the c.1504-9G site functioning as a splicing receptor.

中文翻译：

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ANK1基因新变体c.1504-9G>A的鉴定及其遗传性球形红细胞增多症中内含子保留的机制

目的：本研究的目的是查明两名遗传性球形红细胞增多症儿科患者的致病基因并评估其突变致病性。方法：我们利用全外显子组测序（WES）进行个体分析（病例1）和基于家系的三人组分析（病例1）。情况2）。内含子突变的重要性通过 Minigene 剪接测定得到验证，并得到后续研究的支持体外实验结果：两名先证者均被诊断为遗传性球形红细胞增多症。 WES 发现了一本小说ANK1两名患者中的c.1504-9G＞A突变，导致内含子13的5'端保留7个核苷酸，如Minigene测定所证实的。该变体导致过早终止密码子并产生截短的蛋白质。体外研究表明表达减少ANK1基因.结论：小说ANK1c.1504-9G＞A变体被确定为遗传性球形红细胞增多症的致病因素，其中c.1504-9G位点充当剪接受体。