Chromosomal Haplotypes by Genetic Phasing of Human Families

Roach, Jared C.; Glusman, Gustavo; Hubley, Robert; Montsaroff, Stephen Z.; Holloway, Alisha K.; Mauldin, Denise E.; Srivastava, Deepak; Garg, Vidu; Pollard, Katherine S.; Galas, David J.; Hood, Leroy; Smit, Arian F. A.

doi:10.1016/j.ajhg.2011.07.023

Cited by 63 publications

(70 citation statements)

References 21 publications

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“…In brief, whole-genome sequencing was performed by Complete Genomics using a proprietary paired-end, nanoarray-based sequencing-by-ligation technology [41][42][43] . Seven individuals were selected for whole-genome sequencing, the six core family members (V.4, V.5, V.8, IV.5, IV.6 and IV40) and IV.…”

Section: Methodsmentioning

confidence: 99%

Mutations in STX1B, encoding a presynaptic protein, cause fever-associated epilepsy syndromes

et al. 2014

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Febrile seizures affect 2-4% of all children and have a strong genetic component. Recurrent mutations in three main genes (SCN1A, SCN1B and GABRG2) have been identified that cause febrile seizures with or without epilepsy. Here we report the identification of mutations in STX1B, encoding syntaxin-1B, that are associated with both febrile seizures and epilepsy. Whole-exome sequencing in independent large pedigrees identified cosegregating STX1B mutations predicted to cause an early truncation or an in-frame insertion or deletion. Three additional nonsense or missense mutations and a de novo microdeletion encompassing STX1B were then identified in 449 familial or sporadic cases. Video and local field potential analyses of zebrafish larvae with antisense knockdown of stx1b showed seizure-like behavior and epileptiform discharges that were highly sensitive to increased temperature. Wild-type human syntaxin-1B but not a mutated protein rescued the effects of stx1b knockdown in zebrafish. Our results thus implicate STX1B and the presynaptic release machinery in fever-associated epilepsy syndromes.

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Section: Methodsmentioning

confidence: 99%

Mutations in STX1B, encoding a presynaptic protein, cause fever-associated epilepsy syndromes

et al. 2014

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“…From inheritance, each embryo must be heterozygous at these positions except in rare cases where gene conversion has taken place or where errors are made in the parental genomes. Previous studies (Drmanac et al 2010;Roach et al 2010Roach et al , 2011 using Complete Genomics' sequencing process with a large amount of input DNA suggest that the overall error rate for the parents should be very low and contribute little to this sensitivity calculation. Analysis of the approximately 463,000 loci that met these criteria resulted in a 5.39%-14.39% overall reduction in called heterozygous SNVs (false-negative rate) due to removal by phasing or a lack of sequence coverage in each embryo micro-biopsy (Supplemental Table 5).…”

Section: Assessing Sensitivity and Reducing False-positive Variants Imentioning

confidence: 99%

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

et al. 2015

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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, >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.

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“…These genomes were previously phased at a high quality using familial information 3 . Two phasing libraries were prepared for each member of the trio.…”

Section: Resultsmentioning

confidence: 99%

Whole-genome haplotyping using long reads and statistical methods

et al. 2014

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Rapid growth of sequencing technologies has greatly contributed to increasing our understanding of human genetics. Yet, in spite of this growth, mainstream technologies have been largely unsuccessful in resolving the diploid nature of the human genome. Here we describe statistically aided long read haplotyping (SLRH), a rapid, accurate method based on a simple experimental protocol that requires potentially as little as 30 Gbp of sequencing in addition to a standard (50x coverage) whole-genome analysis for human samples. Using this technology, we phase 99% of single-nucleotide variants in three human genomes into long haplotype blocks of 200 kbp to 1 Mbp in length. As a demonstration of the potential applications of our method, we determine allele-specific methylation patterns in a human genome and identify hundreds of differentially methylated regions that were previously unknown. Such information may offer insight into the mechanisms behind differential gene expression.

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Chromosomal Haplotypes by Genetic Phasing of Human Families

Cited by 63 publications

References 21 publications

Mutations in STX1B, encoding a presynaptic protein, cause fever-associated epilepsy syndromes

Mutations in STX1B, encoding a presynaptic protein, cause fever-associated epilepsy syndromes

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

Whole-genome haplotyping using long reads and statistical methods

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