Semiconductor Sequencing for Preimplantation Genetic Testing for Aneuploidy
Semiconductor Sequencing for Preimplantation Genetic Testing for Aneuploidy
复制标题
用于非整倍体植入前基因检测的半导体测序
DOI：
10.3791/59273
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发表时间：
2019-08-01
影响因子：
1.2
通讯作者：
Choy, Kwong Wai
中科院分区：
文献类型：
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作者：
Gui, Baoheng;Zhang, Yingxin;Choy, Kwong Wai
染色体非整倍体是导致胚胎发育停滞、着床失败或妊娠失败的主要原因之一，在人类胚胎中已有大量文献报道。非整倍体植入前基因检测（PGT-A）是一种通过检测胚胎染色体异常来显着改善生殖结果的基因检测。下一代测序（NGS）为遗传分析提供了高通量和具有成本效益的方法，并已在PGT-A中显示出临床适用性。在这里，我们提出了一种快速和低成本的半导体测序为基础的NGS方法，用于筛选胚胎中的非整倍体。工作流程的第一步是活检胚胎标本的全基因组扩增（WGA），然后构建测序文库，随后在半导体测序系统上进行测序。通常，对于PGT-A应用，可以在每个芯片上加载24个样品并测序，以150个碱基对的平均读取长度产生6000 - 8000万个读取。该方法提供了一种用于进行测序文库的模板扩增和富集的改进方案，使得PGT-A检测具有可重复性、高通量、成本有效和节省时间。这种半导体测序仪的运行时间仅为2-4小时，将从接收样品到出具报告的周转时间缩短为5天。这些优点使该方法成为检测胚胎染色体非整倍性的理想方法，从而有利于其在PGT-A中的广泛应用。
Chromosomal aneuploidy, one of the main causes leading to embryonic development arrest, implantation failure, or pregnancy loss, has been well documented in human embryos. Preimplantation genetic testing for aneuploidy (PGT-A) is a genetic test that significantly improves reproductive outcomes by detecting chromosomal abnormalities of embryos. Next-generation sequencing (NGS) provides a high-throughput and cost-effective approach for genetic analysis and has shown clinical applicability in PGT-A. Here, we present a rapid and low-cost semiconductor sequencing-based NGS method for screening of aneuploidy in embryos. The first step of the workflow is whole genome amplification (WGA) of the biopsied embryo specimen, followed by construction of sequencing library, and subsequent sequencing on the semiconductor sequencing system. Generally, for a PGT-A application, 24 samples can be loaded and sequenced on each chip generating 60-80 million reads at an average read length of 150 base pairs. The method provides a refined protocol for performing template amplification and enrichment of sequencing library, making the PGT-A detection reproducible, high-throughput, cost-efficient, and timesaving. The running time of this semiconductor sequencer is only 2-4 hours, shortening the turnaround time from receiving samples to issuing reports into 5 days. All these advantages make this assay an ideal method to detect chromosomal aneuploidies from embryos and thus, facilitate its wide application in PGT-A.