Application of third-generation sequencing for genetic testing of thalassemia in Guizhou Province, Southwest China

ABSTRACT

Objectives

To explore the application of third-generation sequencing (TGS) for genetic diagnosis and prenatal genetic screening of thalassemia genes.

Methods

Two groups of subjects were enrolled in this study. The first group included 176 subjects with positive hematological phenotypes for thalassemia. Thalassemia-associated genes were detected simultaneously in each sample using both the PacBio TGS platform based on single-molecule real-time (SMRT) technology and the conventional PCR-reverse dot blot (PCR-RDB). Sanger sequencing was used for validation when results were discordant between the two methods. The second group included 53 couples with at least one partner having a positive thalassemia hematological phenotype, and they were screened for homotypic thalassemia variants by TGS, and the risk of pregnancies with babies presenting with severe thalassemia, was assessed.

Results

Of the 176 subjects, 175 had concordant genotypes between the two methods, including 63 normal subjects and 112 α- and/or β-thalassemia gene carriers, with a concordance rate of 99.43%. TGS detected a rare β-thalassemia gene variant −50 (G > A) that was not detected by conventional PCR-RDB. TGS identified seven of the 53 couples as homotypic thalassemia gene carriers, five of whom were at risk of pregnancies with severe thalassemia.

Conclusion

TGS could effectively detect common and rare thalassemia variants with high accuracy and efficiency. This approach would be suitable for prenatal thalassemia genetic screening in areas with high incidence of thalassemia.

KEYWORDS:

• Third-generation sequencing (TGS)
• single-molecule real-time technology (SMRT)
• thalassemia
• Guizhou

Acknowledgements

The authors thank tall individuals for their participation in this study. The authors would like to express their gratitude to EditSprings (https://www.editsprings.cn) for the expert linguistic services provided.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Ethics statement

Informed consent was obtained according to the Declaration of Helsinki.

Data availability statement

All data in this study are shown in the figures and tables.

Additional information

Funding

This work was supported by the [National Natural Science Foundation of China] under Grant [number 81960040]; [Guizhou Provincial Science and Technology Projects] under Grant [numbers 20165670, 20192808, 20191206, 20205011].