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Open AccessArticle

Optimized NGS Approach for Detection of Aneuploidies and Mosaicism in PGT-A and Imbalances in PGT-SR

1
R&D Department, Igenomix, 46980 Valencia, Spain
2
Igenomix Foundation, 46980 Valencia, Spain
3
School of Medicine, University of Valencia/INCLIVA, Valencia 46106, Spain
4
Department of Obstetrics and Gynecology, School of Medicine, Stanford University, Stanford, CA 94305, USA
5
Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX 77030, USA
*
Author to whom correspondence should be addressed.
Carmen M. García-Pascual and Luis Navarro-Sánchez have contributed equally.
Genes 2020, 11(7), 724; https://doi.org/10.3390/genes11070724
Received: 19 May 2020 / Revised: 16 June 2020 / Accepted: 24 June 2020 / Published: 29 June 2020
(This article belongs to the Special Issue EmbryoGenetics)
The detection of chromosomal aneuploidies and mosaicism degree in preimplantation embryos may be essential for achieving pregnancy. The aim of this study was to determine the robustness of diagnosing homogenous and mosaic aneuploidies using a validated algorithm and the minimal resolution for de novo and inherited deletions and duplications (Del/Dup). Two workflows were developed and validated: (a,b) preimplantation genetic testing for uniform whole and segmental aneuploidies, plus mixtures of euploid/aneuploid genomic DNA to develop an algorithm for detecting mosaicism; and (c) preimplantation genetic testing for structural rearrangements for detecting Del/Dup ≥ 6 Mb. Next-generation sequencing (NGS) was performed with automatic library preparation and multiplexing up to 24–96 samples. Specificity and sensitivity for PGT-A were both 100% for whole chromosomes and segmentals. The thresholds stablished for mosaicism were: euploid embryos (<30% aneuploidy), low mosaic (from 30% to <50%), high mosaic (50–70%) or aneuploid (>70%). In the PGT-SR protocol, changes were made to increase the detection level to ≥6 Mb. This is the first study reporting an accurate assessment of semiautomated-NGS protocols using Reproseq on pools of cells. Both protocols allow for the analysis of homogeneous and segmental aneuploidies, different degrees of mosaicism, and small Del/Dup with high sensitivity and specificity. View Full-Text
Keywords: NGS; aneuploidy; mosaicism; segmental; translocations; PGT-A NGS; aneuploidy; mosaicism; segmental; translocations; PGT-A
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  • Externally hosted supplementary file 1
    Doi: 10.5281/zenodo.3834181
    Link: https://zenodo.org/record/3834181#.XsPy_WgzbFQ
    Description: Suppl. Table. 1. Cell lines and their karyotypes used in the validations. In the case of cells carriers of segmental aneuploidies and/or Del/Dup, the size of the fragments was estimated by NGS.
MDPI and ACS Style

García-Pascual, C.M.; Navarro-Sánchez, L.; Navarro, R.; Martínez, L.; Jiménez, J.; Rodrigo, L.; Simón, C.; Rubio, C. Optimized NGS Approach for Detection of Aneuploidies and Mosaicism in PGT-A and Imbalances in PGT-SR. Genes 2020, 11, 724.

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