[Application of single-sperm sequencing in resolving the carrier status of preimplantation genetic testing for chromosomal structural rearrangements in Robertsonian translocations].

Objective: To investigate the application value of single-sperm sequencing in resolving the carrier status of preimplantation genetic testing (PGT) for chromosomal structural rearrangements in Robertsonian translocations.

Methods: Haplotypes were constructed by single-sperm isolation combined with single-sperm sequencing for a patient with 45, XY, der(13; 14)(q10; q10). Twenty single-sperm samples were isolated by mechanical braking and subjected to whole-genome amplification (WGA), and then the Asian Screening Array (ASA) gene chip was used to detect the 183 708 single nucleotide polymorphisms (SNP) of the WGA products. The single sperm associated with the translocation that could be used as haplotype inference was detected by copy number variation (CNV) sequencing, and the chromosomal haplotypes with normal and Robertsonian translocations were inferred. Three biopsy samples of embryonic trophoblast cells were used as the objects. After whole-genome amplification, high-throughput sequencing was employed to determine the status of the translocation chromosome carried by the embryos. The available blastocysts were selected for transfer, and the amniotic fluid samples were taken at 18 weeks of gestation to confirm whether the fetus carried the pathogenic mutation.

Results: A total of 6 037 SNP sites were screened by single-sperm sequencing, and 30 sites selected to distinguish normal and translocation haplotypes. Preimplantation haplotype analysis showed that all the three embryos were euploids without Robertsonian translocation chromosome. Genetic testing of amniotic fluid in the second trimester confirmed that the karyotype of the fetus was 46, XN, carrying no Robertsonian translocation chromosome.

Conclusion: For male carriers of Robertsonian translocation, single sperm sequencing can be used to screen SNP sites to construct haplotypes for distinguishing normal and Robertsonian translocation embryos, and to provide a basis for embryo selection by preimplantation chromosomal structural genetic testing.