PDF(Pubmed)
Abstract：
OBJECTIVE: Various screening techniques have been developed for preimplantation genetic testing for aneuploidy (PGT-A) to reduce implantation failure and miscarriages in women undergoing in vitro fertilisation (IVF) treatment. Among these methods, the Oxford nanopore technology (ONT) has already been tested in several tissues. However, no studies have applied ONT to polar bodies, a cellular material that is less restrictively regulated for PGT-A in some countries.
METHODS: We performed rapid short nanopore sequencing on pooled first and second polar bodies of 102 oocytes from women undergoing IVF treatment to screen for aneuploidy. An automated analysis pipeline was developed with the expectation of three chromatids per chromosome. The results were compared to those obtained by array-based comparative genomic hybridisation (aCGH).
RESULTS: ONT and aCGH were consistent for 96% (98/102) of sample ploidy classification. Of those samples, 36 were classified as euploid, while 62 were classified as aneuploid. The four discordant samples were assessed as euploid using aCGH but classified as aneuploid using ONT. The concordance of the ploidy classification (euploid, gain, or loss) per chromosome was 92.5% (2169 of 2346 of analysed chromosomes) using aCGH and ONT and increased to 97.7% (2113/2162) without the eight samples assessed as highly complex aneuploid using ONT.
CONCLUSIONS: The automated detection of the ploidy classification per chromosome and shorter duplications or deletions depending on the sequencing depth demonstrates an advantage of the ONT method over standard, commercial aCGH methods, which do not consider the presence of three chromatids in pooled polar bodies.
METHODS: We performed rapid short nanopore sequencing on pooled first and second polar bodies of 102 oocytes from women undergoing IVF treatment to screen for aneuploidy. An automated analysis pipeline was developed with the expectation of three chromatids per chromosome. The results were compared to those obtained by array-based comparative genomic hybridisation (aCGH).
RESULTS: ONT and aCGH were consistent for 96% (98/102) of sample ploidy classification. Of those samples, 36 were classified as euploid, while 62 were classified as aneuploid. The four discordant samples were assessed as euploid using aCGH but classified as aneuploid using ONT. The concordance of the ploidy classification (euploid, gain, or loss) per chromosome was 92.5% (2169 of 2346 of analysed chromosomes) using aCGH and ONT and increased to 97.7% (2113/2162) without the eight samples assessed as highly complex aneuploid using ONT.
CONCLUSIONS: The automated detection of the ploidy classification per chromosome and shorter duplications or deletions depending on the sequencing depth demonstrates an advantage of the ONT method over standard, commercial aCGH methods, which do not consider the presence of three chromatids in pooled polar bodies.
摘要：
目的：已经开发了各种筛查技术用于非整倍体（PGT-A）的植入前遗传学检测，以减少接受体外受精（IVF）治疗的女性的植入失败和流产。在这些方法中,牛津纳米孔技术（ONT）已经在几种组织中进行了测试。然而,没有研究将ONT应用于极体，一种细胞材料，在一些国家对PGT-A的监管不太严格。
方法：我们对来自接受IVF治疗的女性的102个卵母细胞的合并的第一和第二极体进行了快速短纳米孔测序，以筛选非整倍性。开发了自动分析流程，期望每个染色体有三个染色单体。将结果与通过基于阵列的比较基因组杂交(aCGH)获得的结果进行比较。
结果：ONT和aCGH对于96％（98/102）的样品倍性分类是一致的。在这些样本中，36个被归类为整倍体，而62个被归类为非整倍体。使用aCGH将四个不一致样品评估为整倍体，但使用ONT分类为非整倍体。倍性分类的一致性(整倍体，增益，或丢失）使用aCGH和ONT的每条染色体为92.5％（分析染色体中的2346个中的2169个），并且在没有使用ONT评估为高度复杂的非整倍体的八个样品的情况下增加到97.7％（2113/2162）。
结论：自动检测每个染色体的倍性分类以及根据测序深度的较短重复或缺失，证明了ONT方法优于标准方法，商业ACGH方法，不考虑在合并的极体中存在三个染色单体。
方法：我们对来自接受IVF治疗的女性的102个卵母细胞的合并的第一和第二极体进行了快速短纳米孔测序，以筛选非整倍性。开发了自动分析流程，期望每个染色体有三个染色单体。将结果与通过基于阵列的比较基因组杂交(aCGH)获得的结果进行比较。
结果：ONT和aCGH对于96％（98/102）的样品倍性分类是一致的。在这些样本中，36个被归类为整倍体，而62个被归类为非整倍体。使用aCGH将四个不一致样品评估为整倍体，但使用ONT分类为非整倍体。倍性分类的一致性(整倍体，增益，或丢失）使用aCGH和ONT的每条染色体为92.5％（分析染色体中的2346个中的2169个），并且在没有使用ONT评估为高度复杂的非整倍体的八个样品的情况下增加到97.7％（2113/2162）。
结论：自动检测每个染色体的倍性分类以及根据测序深度的较短重复或缺失，证明了ONT方法优于标准方法，商业ACGH方法，不考虑在合并的极体中存在三个染色单体。
