Backenroth Daniel, Altarescu Gheona, Zahdeh Fouad, Mann Tzvia, Murik Omer, Renbaum Paul, Segel Reeval, Zeligson Sharon, Hakam-Spector Elinor, Carmi Shai, Zeevi David A
Braun School of Public Health and Community Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel.
PGT Unit, Medical Genetics Institute, Shaare Zedek Medical Center, Jerusalem, Israel.
Sci Rep. 2023 Oct 21;13(1):18036. doi: 10.1038/s41598-023-45292-z.
Recent advances in genomic technologies expand the scope and efficiency of preimplantation genetic testing (PGT). We previously developed Haploseek, a clinically-validated, variant-agnostic comprehensive PGT solution. Haploseek is based on microarray genotyping of the embryo's parents and relatives, combined with low-pass sequencing of the embryos. Here, to increase throughput and versatility, we aimed to develop a sequencing-only implementation of Haploseek. Accordingly, we developed SHaploseek, a universal PGT method to determine genome-wide haplotypes of each embryo based on low-pass (≤ 5x) sequencing of the parents and relative(s) along with ultra-low-pass (0.2-0.4x) sequencing of the embryos. We used SHaploseek to analyze five single lymphoblast cells and 31 embryos. We validated the genome-wide haplotype predictions against either bulk DNA, Haploseek, or, at focal genomic sites, PCR-based PGT results. SHaploseek achieved > 99% concordance with bulk DNA in two families from which single cells were derived from grown-up children. In embryos from 12 PGT families, all of SHaploseek's focal site haplotype predictions were concordant with clinical PCR-based PGT results. Genome-wide, there was > 99% median concordance between Haploseek and SHaploseek's haplotype predictions. Concordance remained high at all assayed sequencing depths ≥ 2x, as well as with only 1ng of parental DNA input. In subtelomeric regions, significantly more haplotype predictions were high-confidence in SHaploseek compared to Haploseek. In summary, SHaploseek constitutes a single-platform, accurate, and cost-effective comprehensive PGT solution.
基因组技术的最新进展扩大了植入前基因检测（PGT）的范围和效率。我们之前开发了Haploseek，这是一种经过临床验证的、不依赖变异类型的综合PGT解决方案。Haploseek基于对胚胎父母及亲属的微阵列基因分型，并结合胚胎的低深度测序。在此，为了提高通量和通用性，我们旨在开发一种仅基于测序的Haploseek实施方案。因此，我们开发了SHaploseek，这是一种通用的PGT方法，可基于对父母及亲属的低深度（≤5x）测序以及对胚胎的超低深度（0.2 - 0.4x）测序来确定每个胚胎的全基因组单倍型。我们使用SHaploseek分析了5个单个淋巴细胞和31个胚胎。我们针对大量DNA、Haploseek或在特定基因组位点的基于PCR的PGT结果验证了全基因组单倍型预测。在两个单细胞来源于成年子女的家庭中，SHaploseek与大量DNA的一致性超过99%。在12个PGT家庭的胚胎中，SHaploseek所有特定位点的单倍型预测均与基于临床PCR的PGT结果一致。在全基因组范围内，Haploseek和SHaploseek的单倍型预测之间的中位数一致性超过99%。在所有检测的测序深度≥2x以及仅输入1ng亲本DNA时，一致性仍然很高。在亚端粒区域，与Haploseek相比，SHaploseek有显著更多的单倍型预测是高可信度的。总之，SHaploseek构成了一个单平台、准确且经济高效的综合PGT解决方案。
