Abstract
Deep targeted sequencing technologies are still not widely used in clinical practice due to the complexity of the methods and their cost. The Molecular Inversion Probes (MIP) technology is cost effective and scalable in the number of targets, however, suffers from low overall performance especially in GC rich regions. In order to improve the MIP performance, we sequenced a large cohort of healthy individuals (n = 4417), with a panel of 616 MIPs, at high depth in duplicates. To improve the previous state-of-the-art statistical model for low variant allele frequency, we selected 4635 potentially positive variants and validated them using amplicon sequencing. Using machine learning prediction tools, we significantly improved precision of 10-56.25% (P < 0.0004) to detect variants with VAF > 0.005. We further developed biochemically modified MIP protocol and improved its turn-around-time to ∼4 h. Our new biochemistry significantly improved uniformity, GC-Rich regions coverage, and enabled 95% on target reads in a large MIP panel of 8349 genomic targets. Overall, we demonstrate an enhancement of the MIP targeted sequencing approach in both detection of low frequency variants and in other key parameters, paving its way to become an ultrafast cost-effective research and clinical diagnostic tool.
Original language | English |
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Article number | lqab125 |
Journal | NAR Genomics and Bioinformatics |
Volume | 4 |
Issue number | 1 |
DOIs | |
Publication status | Published - 1 Mar 2022 |
Funding
Biological samples from Mobilized PBSC autologous transplant products were generously provided by Dr Mark Minden. L.S. is the incumbent of The Ruth and Louis Leland career development chair. FUNDING EU horizon 2020 grant project MAMLE [714731]; LLS and rising tide foundation [RTF6005-19]; ISF-NSFC [2427/18]; ISF-IPMP-Israel Precision Medicine Program [3165/19, BIRAX 713023]; Ernest and Bonnie Beutler Research Program of Excellence in Genomic Medicine (to L.I.S.); L.I.S. is an incumbent of the Ruth and Louis Leland career development chair; N.K. is an incumbent of the Applebaum Foundation Research Fellow Chair; Sagol Institute for Longevity Research; Barry and Eleanore Reznik Family Cancer Research Fund; Steven B. Rubenstein Research Fund for Leukemia and Other Blood Disorders; Rising Tide Foundation; Applebaum Foundation; Princess Margaret Cancer Foundation (to D.K., in part).
All Science Journal Classification (ASJC) codes
- Structural Biology
- Molecular Biology
- Genetics
- Computer Science Applications
- Applied Mathematics