Application of next-generation sequencing for 24-chromosome aneuploidy screening of human preimplantation embryos

Abstract

BackgroundAneuploidy is a leading cause of repeat implantation failure and recurrent miscarriages. Preimplantation genetic screening (PGS) enables the assessment of the numeral and structural chromosomal errors of embryos before transfer in patients undergoing in vitro fertilization. Array comparative genomic hybridization (aCGH) has been demonstrated to be an accurate PGS method and in present thought to be the gold standard, but new technologies, such as next-generation sequencing (NGS), continue to emerge. Validation of the new comprehensive NGS-based 24-chromosome aneuploidy screening technology is still needed to determine the preclinical accuracy before it might be considered as an alternative method for human PGS.ResultsIn the present study, 43 human trophectoderm (TE) biopsy samples and 5 cytogenetically characterized cell lines (Coriell Cell Repositories) were tested. The same whole genome amplified product of each sample was blindly assessed with Veriseq NGS and Agilent aCGH to identify the aneuploidy status. The result showed that the NGS identified all abnormalities identified in aCGH including the numeral chromosomal abnormalities (again or loss) in the embryo samples and the structural (partial deletion and duplication) in the Coriell cell lines. Both technologies can identify a segmental imbalance as small as 1.8 Mb in size. Among the 41 TE samples with abnormal karyotypes in this study, eight (19.5 %) samples presented as multiple chromosome abnormalities. The abnormalities occurred to almost all chromosomes, except chromosome 6, 7, 17 and Y chromosome.ConclusionsGiven its reliability and high level of consistency with an established aCGH methodology, NGS has demonstrated a robust high-throughput methodology ready for extensive clinical application in reproductive medicine, with potential advantages of reduced costs and enhanced precision. Then, a randomized controlled clinical trial confirming its clinical effectiveness is advisable to obtain a larger sequencing dataset and more evidence for the extensive use of NGS-based PGS.

Recent advances in preimplantation genetic diagnosis and screening

“…Many studies also showed that NGS has become an efficient and robust technology for PGD/PGS [67][68][69]. For example, one study reported 24 healthy births after NGS testing [70] and another one resulting in a healthy birth to the parents carrying Robertsonian translocation after NGS-based PGD [71].…”

Assessment of pre‐implantation genetic testing for embryo aneuploidies: A SWOT analysis

“…On the other hand, qPCR does not use WGA, can identify aneuploidies in a rapid fashion but has lower genomic coverage, is not able to distinguish small deletions and duplications and cannot detect structural chromosome aberrations or mosaicisms . NGS is the newest platform for PGT‐A that permits to reduce DNA sequencing cost, increases number of samples that can be simultaneously sequenced, detects unbalanced translocations, partial aneuploidies and enhances the detection of mosaicisms . The higher rate of mosaicism detected by NGS is probably explained by a superior sensitivity of this method for detecting minor lines in mixed cell populations compared with aCGH .…”

“…23 NGS is the newest platform for PGT-A that permits to reduce DNA sequencing cost, increases number of samples that can be simultaneously sequenced, detects unbalanced translocations, partial aneuploidies and enhances the detection of mosaicisms. 24,25 The higher rate of mosaicism detected by NGS is probably explained by a superior sensitivity of this method for detecting minor lines in mixed cell populations compared with aCGH. 26 Even so, depending on the depth of sequencing and the specific NGS platform used, the sensitivity for detecting cytogenetically distinct subpopulations of cells varies as NGS strategies change.…”

Preimplantation genetic testing for aneuploidy: what technology should you use and what are the differences?

“…Next generation sequencing (NGS) is a technology that requires optimized DNA amplification to reduce the introduction of artifacts during the amplification process [49,50]. Following the DNA amplification, artifacts can be identified and removed by bioinformatics software.…”