Clinical application of next generation sequencing-based haplotype linkage analysis in the preimplantation genetic testing for germline mosaicisms

Orphanet Journal of Rare Diseases

Research

Yu Fu¹  Rong Li²  Yali Wang²  Jiafu Pan²  Chenhui Ding²  Dongjia Chen²  Yanhong Zeng²  Jing Wang²  Yuliang Liu²  Bing Cai²  Han Zhang²  Jing Guo²  Yan Xu²  Canquan Zhou³  Xiaoting Shen³ 

[1] The First Affiliated Hospital of Hainan Medical University, Hainan Medical University, 570102, Haikou, China;The First Affiliated Hospital, Sun Yat-Sen University, 510080, Guangzhou, China;Guangdong Provincial Key Laboratory of Reproductive Medicine, 510080, Guangzhou, China;The First Affiliated Hospital, Sun Yat-Sen University, 510080, Guangzhou, China;Guangdong Provincial Key Laboratory of Reproductive Medicine, 510080, Guangzhou, China;Reproductive Medicine Center, The First Affiliated Hospital, Sun Yat-sen University, 510080, Guangzhou, China;

关键词: Germline mosaicism;    Next generation sequencing;    Single nucleotide polymorphism;    Preimplantation genetic testing;    Haplotype linkage analysis;   

DOI  :  10.1186/s13023-023-02736-z

 received in 2022-03-29, accepted in 2023-05-18,  发布年份 2023

来源: Springer

【 摘 要 】

BackgroundPreimplantation genetic testing (PGT) for monogenic disorders (PGT-M) for germline mosaicism was previously highly dependent on polymerase chain reaction (PCR)-based directed mutation detection combined with linkage analysis of short tandem repeats (STRs). However, the number of STRs is usually limited. In addition, designing suitable probes and optimizing the reaction conditions for multiplex PCR are time-consuming and laborious. Here, we evaluated the effectiveness of next generation sequencing (NGS)-based haplotype linkage analysis in PGT of germline mosaicism.MethodsPGT-M with NGS-based haplotype linkage analysis was performed for two families with maternal germline mosaicism for an X-linked Duchenne muscular dystrophy (DMD) mutation (del exon 45–50) or an autosomal TSC1 mutation (c.2074C > T). Trophectoderm biopsy and multiple displacement amplification (MDA) were performed for a total of nine blastocysts. NGS and Sanger sequencing were performed in genomic DNA of family members and embryonic MDA products to detect DMD deletion and TSC1 mutation, respectively. Single nucleotide polymorphism (SNP) sites closely linked to pathogenic mutations were detected with NGS and served in haplotype linkage analysis. NGS-based aneuploidy screening was performed for all embryos to reduce the risk of pregnancy loss.ResultsAll nine blastocytes showed conclusive PGT results. Each family underwent one or two frozen-thawed embryo transfer cycles to obtain a clinical pregnancy, and the prenatal diagnosis showed that the fetus was genotypically normal and euploid for both families.ConclusionsNGS-SNP could effectively realize PGT for germline mosaicism. Compared with PCR-based methods, the NGS-SNP method with increased polymorphic informative markers can achieve a greater diagnostic accuracy. Further studies are warranted to verify the effectiveness of NGS-based PGT of germline mosaicism cases in the absence of surviving offsprings.

【 授权许可】

CC BY   
© The Author(s) 2023

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