Alan Berkeley

Bahram G. Kermani

Birgit Crain

Brock A. Peters

Daniel M. Hayden

Mark A. McElwain

Misha R. Agarwal

Natali Gulbahce

Oleg Alferov

Radoje Drmanac

Rebecca Yu Zhang

Renata Prates

Rick Tearle

Santiago Munné

Y. Tom Tang

Currently, the methods available for preimplantation genetic diagnosis (PGD) of in vitro fertilized (IVF) embryos do not detect de novo single-nucleotide and short indel mutations, which have been shown to cause a large fraction of genetic diseases. Detection of all these types of mutations requires whole-genome sequencing (WGS). In this study, advanced massively parallel WGS was performed on three 5- to 10-cell biopsies from two blastocyst-stage embryos. Both parents and paternal grandparents were also analyzed to allow for accurate measurements of false-positive and false-negative error rates. Overall, &gt;95% of each genome was called. In the embryos, experimentally derived haplotypes and barcoded read data were used to detect and phase up to 82% of de novo single base mutations with a false-positive rate of about one error per Gb, resulting in fewer than 10 such errors per embryo. This represents a ∼100-fold lower error rate than previously published from 10 cells, and it is the first demonstration that advanced WGS can be used to accurately identify these de novo mutations in spite of the thousands of false-positive errors introduced by the extensive DNA amplification required for deep sequencing. Using haplotype information, we also demonstrate how small de novo deletions could be detected. These results suggest that phased WGS using barcoded DNA could be used in the future as part of the PGD process to maximize comprehensiveness in detecting disease-causing mutations and to reduce the incidence of genetic diseases.</jats:p

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Genome Research

Detection and phasing of single base de novo mutations in biopsies from human in vitro fertilized embryos by advanced whole-genome sequencing

Detection and phasing of single base de novo mutations in biopsies from human in vitro fertilized embryos by advanced whole-genome sequencing

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