A reference data set of 5.4 million phased human variants validated by genetic inheritance from sequencing a three-generation 17-member pedigree

Eberle, Michael A.; Fritzilas, Epameinondas; Krusche, Peter; Källberg, Morten; Moore, Benjamin L.; Bekritsky, Mitchell A.; Iqbal, Zamin; Chuang, Han‐Yu; Humphray, Sean; Halpern, Aaron L.; Kruglyak, Semyon; Margulies, Elliott H.; McVean, Gil; Bentley, David

doi:10.1101/gr.210500.116

Cited by 362 publications

(392 citation statements)

References 36 publications

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“…The 50× paired-end and PCR-free Illumina sequencing data for the trio composed of the individuals NA12891 (father), NA12892 (mother), and daughter (NA12878) were obtained from the European Nucleotide Archive (accession number ERP001960). The data were generated as part of the Illumina Platinum Genomes project 46 , but the data for individual NA12878 were also used to generate the 1000 Genomes Project structural variant calls 7 . The sample input to BayesTyper was obtained by counting the occurrences of all 55-mers in the sequencing data from the three individuals using KMC2 with counting of singletons enabled.…”

Section: Methodsmentioning

confidence: 99%

Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

Maretty

Jensen

Petersen

et al. 2017

Nature

138

149

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Hundreds of thousands of human genomes are now being sequenced to characterize genetic variation and use this information to augment association mapping studies of complex disorders and other phenotypic traits 1-4 . Genetic variation is identified mainly by mapping short reads to the reference genome or by performing local assembly 2,5-7 . However, these approaches are biased against discovery of structural variants and variation in the more complex parts of the genome. Hence, large-scale de novo assembly is needed. Here we show that it is possible to construct excellent de novo assemblies from high-coverage sequencing with mate-pair libraries extending up to 20 kilobases. We report de novo assemblies of 150 individuals (50 trios) from the GenomeDenmark project. The quality of these assemblies is similar to those obtained using the more expensive long-read technology 4,8-13 . We use the assemblies to identify a rich set of structural variants including many novel insertions and demonstrate how this variant catalogue enables further deciphering of known association mapping signals. We leverage the assemblies to provide 100 completely resolved major histocompatibility complex haplotypes and to resolve major parts of the Y chromosome. Our study provides a regional reference genome that we expect will improve the power of future association mapping studies and hence pave the way for precision medicine initiatives, which now are being launched in many countries including Denmark.Using a combination of high-depth (average 78× ) Illumina pairedend and mate-pair libraries, we applied Allpaths-LG 14 to create de novo assemblies of high quality and coverage for each of the 150 individuals with a median scaffold N50 of ~ 21 megabases (Mb; maximum ~ 30 Mb) (Supplementary Table 1). The 100 largest scaffolds in each of the 140 best assemblies typically covered more than 75% (median 77%, Extended Data Fig. 1a) of the genome, with the largest scaffolds exceeding 110 Mb in size (Supplementary Table 1). To evaluate the accuracy of the assemblies, we subsequently aligned the scaffolds for each individual to the human reference genome (GRCh38) 15 . Figure 1 shows an example individual where the euchromatic part of each chromosome was almost completely covered by a few large scaffolds and in several cases scaffolds covered almost entire chromosome arms. Only rarely did we find that large scaffolds break and align to more than one chromosome (Extended Data Fig. 1b), suggesting that even the largest scaffolds are seldom chimaeric. We also compared our de novo assemblies with a published long-read assembly based on BioNano mapping and PacBio sequencing 16 . Extended Data Figs 2a and 3 show that this assembly was less complete than our assemblies, but with similar scaffold lengths. The long-read assembly had 5.38% missing data compared with our median of 4.25% (Extended Data Fig. 3a), but the missing data in our assemblies were found in smaller gaps (Extended Data Fig. 3b, c), and the median contig length was therefore much smaller th...

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Section: Methodsmentioning

confidence: 99%

Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

Maretty

Jensen

Petersen

et al. 2017

Nature

138

149

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“…In all our analyses, the parameters p and s were set to 0.97 and 5. The values were chosen to maximize Mendelian consistency of genotype calls in Platinum Genome pedigree samples (Eberle et al 2017) on an unrelated set of repeats.…”

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confidence: 99%

Detection of long repeat expansions from PCR-free whole-genome sequence data

et al. 2017

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Identifying large expansions of short tandem repeats (STRs), such as those that cause amyotrophic lateral sclerosis (ALS) and fragile X syndrome, is challenging for short-read whole-genome sequencing (WGS) data. A solution to this problem is an important step toward integrating WGS into precision medicine. We developed a software tool called ExpansionHunter that, using PCR-free WGS short-read data, can genotype repeats at the locus of interest, even if the expanded repeat is larger than the read length. We applied our algorithm to WGS data from 3001 ALS patients who have been tested for the presence of the C9orf72 repeat expansion with repeat-primed PCR (RP-PCR). Compared against this truth data, ExpansionHunter correctly classified all (212/212, 95% CI [0.98, 1.00]) of the expanded samples as either expansions (208) or potential expansions (4). Additionally, 99.9% (2786/2789, 95% CI [0.997, 1.00]) of the wild-type samples were correctly classified as wild type by this method with the remaining three samples identified as possible expansions. We further applied our algorithm to a set of 152 samples in which every sample had one of eight different pathogenic repeat expansions, including those associated with fragile X syndrome, Friedreich's ataxia, and Huntington's disease, and correctly flagged all but one of the known repeat expansions. Thus, ExpansionHunter can be used to accurately detect known pathogenic repeat expansions and provides researchers with a tool that can be used to identify new pathogenic repeat expansions.

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“…Overall, the significant lack of mosaic aneuploidy events in samples from blood (p= 3.08x10 -34 , Binomial Test) and the lack of meiotic events, together with the enrichment of trisomy 12, which is the most common cytogenetic abnormality in chronic B lymphocytic leukemia 28 , combine to suggest that these mosaic aneuploidies arose during cell culture and were either neutral in effect or promoted positive selection for these transformed B lymphocytes. One of the samples in which segmental LOH for distal chromosome 11 was identified was GM12889, for which whole genome sequencing (WGS) to a mean ~50X coverage has been performed to define high-confidence, "platinum" variants 29 . We downloaded those WGS data and re-ran MADSEQ, again predicting the LOH of a 19.3 Mb region, estimated to be present in over 50% of the cells (Figure 2).…”

Section: Application Of Madseq To Sequencing Datamentioning

confidence: 99%

“…Picard (v 1.119) was used to mark duplicates and GATK (v 3.4-46) 25 was used for indel realignment and base recalibration following best practices [27][28][29] .…”

Section: Design Of Multi-ethnic Targeted Locimentioning

confidence: 99%

Mosaic chromosomal aneuploidy detection by sequencing (MAD-seq)

Kong

Berko

Marcketta

et al. 2017

Preprint

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Current approaches to detect and characterize mosaic chromosomal aneuploidy are limited by sensitivity, efficiency, cost or the need to culture cells. We describe a combination of a new sequencing-based assay and a novel analytical approach that allows low levels of mosaicism for chromosomal aneuploidy to be detected, assigned to a meiotic or mitotic origin, and quantified as a proportion of the cells in the sample. We show results from a multi-ethnic assay design that is suitable for populations of diverse racial and ethnic origins, and how the MADSEQ analytical approach applied to exome sequencing data reveals unrecognized aneuploidy in 1000 Genomes samples and cell lines from public repositories. We have made the assay design and analytical software open for unrestricted use, with the goal that it can be applied in clinical samples to allow new insights into the unrecognized prevalence of mosaic chromosomal aneuploidy and its phenotypic associations.

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A reference data set of 5.4 million phased human variants validated by genetic inheritance from sequencing a three-generation 17-member pedigree

Cited by 362 publications

References 36 publications

Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

Detection of long repeat expansions from PCR-free whole-genome sequence data

Mosaic chromosomal aneuploidy detection by sequencing (MAD-seq)

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