A comprehensive preimplantation genetic testing approach for SEA-type α-thalassemia by fluorescent Gap-polymerase chain reaction combined with haplotype analysis

Abstract

Abstract Purpose This study aims to evaluate the feasibility and necessity of using fluorescence Gap-polymerase chain reaction (PCR) combined with haplotype analysis in preimplantation genetic testing (PGT) for SEA-type α-thalassemia.Methods Twenty-six PGT biopsy cycles were performed in 25 families from June 2021 to February 2022. All couples were carriers of the SEA-type α-thalassemia. Fluorescent Gap-PCR was employed for fragment deletion detection. Subsequently, according to the PCR results, reference embryos were identified to establish haplotype using single nucleotide polymorphic (SNP) array, while aneuploidy was screened simultaneously. In the cases that PCR results were inconsistent with the haplotype results, the reasons were investigated, either by re-test of the biopsied samples or re-biopsy of the embryo.Results Among 172 embryos, 162 had a consistent result tested by both methods, leading to a consistency rate of 94.2%. Ten embryos had inconsistent results, which were mainly due to chromosome 16 aneuploidy (n = 7), allele drop-out (ADO) in Gap-PCR (n = 2), or incorrectly haplotype due to poor sample amplification quality (n = 1). Clinical pregnancy rate of each frozen-thawed embryo transfer (FET) was 57.7% (15/26). Six families underwent prenatal diagnosis, which confirmed the PGT results.Conclusions Fluorescent Gap-PCR combined with haplotype analysis is feasible and necessary for SEA-type α-thalassemia PGT.