Application of an optimized non-invasive prenatal testing for thalassemia based on change of haplotype doses

Patients affected by monogenic diseases impose a substantial burden on both themselves and their families. The primary preventive measure, i.e., invasive prenatal diagnosis, carries a risk of miscarriage and cannot be performed early in pregnancy. Hence, there is a need for non-invasive prenatal testing (NIPT) for monogenic diseases. By utilizing enriched cell-free fetal DNA (cffDNA) from maternal plasma, we refine the NIPT method, which combines targeted region capture technology, haplotyping, and analysis of informative site frequency. We apply this method to 93 clinical families at genetic risk for thalassemia, encompassing various genetic variant types, to establish a workflow and evaluate its efficiency. Our approach requires only 3 ng of DNA input to generate 0.1 Gb informative target genomic data and leverages a minimum of 3% cffDNA. This method has a 98.16% success rate and 100% concordance with conventional invasive methods. Furthermore, we demonstrate the ability to analyze fetal genotypes as early as eight weeks of gestation. This study establishes an optimized NIPT method for the early detection of various thalassemia disorders during pregnancy. This technique demonstrates high accuracy and potential for clinical application in prenatal diagnosis.