Center for Reproductive Medicine, Department of Obstetrics and Gynecology, Peking University Third Hospital, No. 49 Hua Yuan Bei Road, Hai Dian District, Beijing, 100191, China.
National Clinical Center for Obstetrics and Gynecology, Beijing, China.
J Assist Reprod Genet. 2021 Aug;38(8):1979-1986. doi: 10.1007/s10815-021-02126-z. Epub 2021 Mar 15.
To determine whether next-generation sequencing (NGS) could be used to directly detect different mutations of Duchenne muscular dystrophy (DMD) during preimplantation genetic testing (PGT).
From Sep. 2016 to Aug. 2018, a total of six couples participated in this study. Four cases carried DMD exon deletions and two carried exon duplications. Trophectoderm cells were biopsied at day 5 or 6 and NGS was used in the genetic testing of the biopsied cells after whole-genome amplification. We developed a new method-DIRected Embryonic Cell Testing of Exon Deletion/Duplication (DIRECTED) to directly detect the single-gene mutation by NGS. Linage analysis based on single-nucleotide polymorphism (SNP) was used to validate the results from DIRECTED.
In the four deletion cases, DIRECTED was used to detect DMD exon deletion in 16 biopsied embryos. All DIRECTED results were consistent with linkage analysis, indicating this method was reliable in detecting deletions around 1 Mb. In the two cases carrying exon duplications, no blastocyst was obtained for biopsy. Nonetheless, preliminary experiment results suggested that DIRECTED could also be used for direct detection of exon duplications in embryos.
Exon deletions or duplications in DMD of preimplantation embryos could be detected directly by NGS-based methods during PGT.
确定下一代测序（NGS）是否可用于在胚胎植入前遗传学检测（PGT）中直接检测杜氏肌营养不良症（DMD）的不同突变。
2016 年 9 月至 2018 年 8 月，共有 6 对夫妇参与了这项研究。4 例携带 DMD 外显子缺失，2 例携带外显子重复。在第 5 或第 6 天对滋养外胚层细胞进行活检，并在全基因组扩增后对活检细胞进行 NGS 遗传检测。我们开发了一种新方法-DIRected Embryonic Cell Testing of Exon Deletion/Duplication（DIRECTED），通过 NGS 直接检测单基因突变。基于单核苷酸多态性（SNP）的谱系分析用于验证 DIRECTED 的结果。
在 4 例缺失病例中，使用 DIRECTED 检测了 16 个活检胚胎中的 DMD 外显子缺失。所有 DIRECTED 结果均与连锁分析一致，表明该方法在检测 1Mb 左右的缺失时可靠。在携带外显子重复的 2 例病例中，未获得活检的囊胚。尽管如此，初步实验结果表明，DIRECTED 也可用于直接检测胚胎中外显子重复。
PGT 期间，可通过基于 NGS 的方法直接检测胚胎中 DMD 的外显子缺失或重复。
