Application of single sperm sequencing for preimplantation genetic testing of male patient with achondroplasia / 中华医学遗传学杂志

ABSTRACT

OBJECTIVE@#To assess the value of single sperm sequencing in preimplantation genetic diagnosis.@*METHODS@#A male patient with achondroplasia due to a de novo FGFR3 variant was subjected to single sperm isolation and sequencing. Twenty single sperm samples were isolated by mechanical immobilization, and their whole genome was amplified. PCR primers were designed for the variant site and 25 flanking single nucleotide polymorphism (SNP) loci, and the PCR products were sequenced to determine the chromosomal haplotype which did not harbor the pathogenic variant. Biopsy samples of 12 embryonic trophoblasts were taken. Following whole genome amplification, high-throughput sequencing was carried out to detect the carrier status of the embryos. Wild type blastocysts were selected for transplantation. Amniotic fluid samples were taken at 19 weeks of gestation to confirm the status of the fetus.@*RESULTS@#Eight SNP were selected by single sperm sequencing, with which the haplotypes were successfully constructed. Preimplantation genetic testing indicated that 5 embryos have carried the pathogenic variant and 7 did not. Testing of amniotic fluid sample during the second trimester of pregnancy confirmed that the fetus did not carry the FGFR3 gene c.1138G>A variant.@*CONCLUSION@#For male patients carrying de novo pathogenic variants, SNP sites can be selected through single sperm sequencing, and haplotypes can be constructed by linkage analysis for preimplantation genetic diagnosis.