Targeted Next-Generation Sequencing Reveals a Large Novel β-Thalassemia Deletion that Removes the Entire HBB Gene

β-Thalassemia (β-thal) is one of the most common monogenic recessive inherited diseases worldwide. The mutation spectrum of β-thal has been increasingly broadened by various genetic testing methods. The discovery and identification of novel and rare pathogenic thalassemia variants enable better disease prevention, especially in high prevalence regions. In this study, a Chinese thalassemia family with an unclear etiology was recruited to the Thalassemia Screening Program. Blood samples collected from them were primarily screened by hematology analysis and clinical routine genetic screening. Subsequently, targeted next-generation sequencing (NGS) and Sanger sequencing were performed to find and identify a novel deletion variant. The deletion, discovered by targeted NGS, was validated through real-time quantitative polymerase chain reaction (qPCR). First, a large novel β-thal deletion (3488 bp) related to a high Hb F level, NC_000011.9: g.5245533_5249020del (Chongqing deletion) (GRCh37/hg19), was found and identified in the proband and her mother. The deletion removed the entire β-globin gene and led to absent β-globin (β0). We then validated this large novel deletion in the proband and her mother by qPCR. We first discovered and identified a large novel β-thal deletion related to elevated Hb F level, it helps broaden the spectrum of pathogenic mutants that may cause β-thal intermedia (β-TI) or β-thal major (β-TM), paving the way for effective thalassemia screening. Next-generation sequencing has the potential of finding rare and novel thalassemia mutants.β地中海贫血（β-thal）是全球范围内最为常见的单基因隐性遗传病之一。各类基因检测技术持续拓宽了β地中海贫血的突变谱系。新型、罕见致病性地中海贫血变异体的发现与鉴定，可助力疾病防控工作的优化，在疾病高发地区尤为如此。本研究招募了一个病因未明的中国地中海贫血家系，纳入地中海贫血筛查项目。研究人员首先对采集自该家系成员的血液样本开展血液学分析与临床常规基因筛查，完成初步筛选；随后通过靶向二代测序（next-generation sequencing，NGS）与桑格测序（Sanger sequencing），寻找并鉴定新型缺失变异。研究人员通过靶向NGS发现的该缺失变异，经实时定量聚合酶链反应（qPCR）验证。首先，在先证者及其母亲体内检出并鉴定出一种与高胎儿血红蛋白（Hb F）水平相关的新型大片段β地中海贫血缺失变异（长度3488 bp），其基因组坐标为NC_000011.9: g.5245533_5249020del（重庆缺失），对应GRCh37/hg19参考基因组。该缺失变异可完全敲除β珠蛋白基因，导致β珠蛋白完全缺失（β0型）。研究人员随后通过qPCR对先证者及其母亲体内的该新型大片段缺失变异进行了验证。本研究首次发现并鉴定出一种与胎儿血红蛋白水平升高相关的新型大片段β地中海贫血缺失变异，该发现有助于拓宽可导致中间型β地中海贫血（β-thal intermedia，β-TI）或重型β地中海贫血（β-thal major，β-TM）的致病性突变谱，为高效的地中海贫血筛查奠定基础。二代测序技术具备发现罕见、新型地中海贫血突变体的应用潜力。