Identification of a novel 107 kb deletion in the alpha-globin gene cluster using third-generation sequencing

Objective: To describe the characterization of a novel deletion causing α-thalassemia.

Methods: The proband, a 30-year-old female, displayed mild anemia from thalassemia screening. Gap-PCR was used to detect the four common deletional α-thalassemia, and a PCR-reverse dot blot was performed for the three point mutations of the α-globin gene. Multiplex ligation-dependent probe amplification (MLPA) was used to query possible breakpoints of a potential novel deletion. Third-generation sequencing (TGS) was used to identify the novel deletion after the MLPA failed. Gap-PCR and Sanger sequencing were validated for the breakpoint.

Results: No abnormal results were detected by Gap-PCR and PCR-reverse dot blot. MLPA only showed a large deletion upstream of the HBZ-1 probe, but the scope could not be determined. However, a novel 107 kb deletion at the α-globin gene was discovered by the TGS. The Gap-PCR products with the specific breakpoint fragment of the 107 kb deletion were confirmed by Sanger sequencing.

Conclusions: A 107 kb deletion causing α-thalassemia was the first reported worldwide. TGS played an important role in this study and can be recommended as a reliable tool for rare or potential deletions in thalassemia.