Affected-embryo-based SNP haplotyping with NGS for the preimplantation genetic testing of Marfan syndrome

Marfan syndrome (MFS), an autosomal dominant heritable disease of the connective tissue, is characterized by broad clinical manifestations in the musculoskeletal, cardiovascular, pulmonary, and ocular systems. In this study, a male patient with MFS caused by a heterozygous mutation NM_000138.5(FBN1):c.6037 + 2 T > C in the fibrillin 1 gene (FBN1) underwent preimplantation genetic testing (PGT) by using affected-embryo-based single nucleotide polymorphism (SNP) haplotyping. Multiple displacement amplification was used for whole genome amplification of biopsied trophectoderm cells after controlled ovarian stimulation. Sanger sequencing and next-generation sequencing (NGS) were used to detect the state of FBN1 mutation. A total of 14 blastocysts formed after intracytoplasmic sperm injection were biopsied. After NGS, 60 informative polymorphic SNP markers located upstream and downstream of the FBN1 gene and its pathogenic mutation site were linked to individual alleles. Sanger sequencing further confirmed that 8 blastocysts carried the mutation NM_000138.5(FBN1):c.6037 + 2 T > C, while 6 did not. Four of the non-carriers were euploid verified by copy number variation results. A female infant without MFS was born at 37 weeks gestation after a subsequent frozen embryo transfer. In conclusion, the successful case indicates that SNP haplotyping using sibling embryos as a reference is applicable to PGT in monogenetic diseases.