Application of third-generation sequencing technology in preimplantation genetic testing of embryos in a family with ATR-X syndrome
Objective To explore the clinical application value of third-generation sequencing technology in preimplantation genetic testing(PGT)of an ATR-X syndrome pedigree with chromosomal microduplication.Methods The study selected a pedigree with a suspected ATR-X syndrome child at Assisted Reproductive Center of Jiangxi Maternal and Child Health Hospital in October 2022.After chromosome copy number variation sequencing(CNV Seq)detection,it was found that the female carried a 550 kb heterozygous microrepeat with unclear clinical significance in the Xq21.1 region,which involved partial sequences of the ATRX gene.The third generation long read sequencing technology was used to detect the female genome sequence,determine the physical location of the insertion of the above repeat into the genome,clarify the pathogenicity of the repeat,and obtain a single nucleotide polymorphism(SNP)haplotype linked to the above micro repeat inheritance.After the couple's full informed consent,PGT was performed to assist pregnancy.One haploid embryo without pathogenic microduplication was selected for transfer.To verify the consistency with PGT test results,amniocentesis prenatal diagnosis was performed in the second trimester after successful pregnancy,and the fetus was followed up after birth.Results The results of the third generation long read sequencing and Sanger sequencing verification showed that the Xq21.1 microrepeat carried by the female was inserted into the genome chrX:76804463-76804464(GRCh37/hg19),which is an intra tandem repeat of the ATRX gene,and it is predicted that it may cause damage to the normal function of the ATRX protein.After one cycle of PGT treatment,27 oocytes were obtained and 13 blastocysts were successfully developed after intracytoplasmic sperm injection(ICSI).Through genetic testing,it was found that two blastocysts were haploid embryos without carrying the aforementioned pathogenic microduplication.After thawing and transferring one of the blastocysts,the pregnancy was achieved,and the prenatal diagnosis results of amniocentesis in the second trimester were consistent with the PGT results.In November 2023,at 39+5 weeks of pregnancy,a female live baby was delivered by natural delivery,and she is in good health.Conclusion The third-generation sequencing technology has significant advantages in PGT detection of clinically ambiguous microreplicates with functional deficiency due to its long read length characteristics.It can not only determine the location of microreplicates inserted into the genome and determine their pathogenicity,but also obtain SNP haplotypes that are linked to the target mutation,thus preparing for subsequent embryo detection.
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