With the rapid advancement of sequencing technology, third-generation sequencing, particularly single-molecule real-time sequencing and nanopore sequencing, has opened new avenues for the precise prevention and control of genetic diseases. It offers distinct advantages, including long-read lengths, minimal requirement for extensive polymerase chain reaction amplification, and real-time sequencing capability. This narrative review summarizes and critically evaluates recent advances in the application and latest progress of third-generation sequencing in thalassemia. It focuses on the technical advantages and application status of this technology in detecting complex structural variations, large-scale carrier screening, prenatal diagnosis, and preimplantation genetic testing. The review further analyzes the considerable potential of this technology in improving diagnostic accuracy, shortening detection cycles, and reducing technical costs. Additionally, it discusses the challenges currently faced in data interpretation, standardized processes, and clinical translation. Finally, this review provides insights into future development directions, with the aim of providing new technical perspectives and a theoretical basis for optimizing the prevention and control strategies of thalassemia.