A recurrence based approach for validating structural variation using long-read sequencing technology

Zhao, Xuefang; Weber, Alexandra M.; Mills, Ryan E.

doi:10.1101/105817

Cited by 13 publications

(16 citation statements)

References 20 publications

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“…The PacBio genome assembly-based approach overlooked this insertion, as it fell within a 'reference L1 rich' region ( Fig.4d ). Upon further inspection, this event was supported as a 403bp heterozygous L1Hs insertion by the existence of significant retrotransposition hallmarks, as well as recurrence (dot) plots 29,66 ( Fig.4e , see Methods ). This insertion also shares a high sequence identity with a nearby reference L1PA11 element ( Supplementary Table 7 ).…”

Section: Cas9 Enrichment and Nanopore Sequencing Captures Non-referenmentioning

confidence: 87%

Cas9 targeted enrichment of mobile elements using nanopore sequencing

McDonald

Zhou

Castro

et al. 2021

Preprint

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Mobile element insertions (MEIs) are highly repetitive genomic sequences that contribute to inter- and intra-individual genetic variation and can lead to genetic disorders. Targeted and whole-genome approaches using short-read sequencing have been developed to identify reference and non-reference MEIs; however, the read length hampers detection of these elements in complex genomic regions. Here, we pair Cas9 targeted nanopore sequencing with computational methodologies to capture active MEIs in human genomes. We demonstrate parallel enrichment for distinct classes of MEIs, averaging 44% of reads on targeted signals. We show an individual flow cell can recover a remarkable fraction of MEIs (97% L1Hs, 93% AluYb, 51% AluYa, 99% SVA_F, and 65% SVA_E). We identify twenty-one non-reference MEIs in GM12878 overlooked by modern, long-read analysis pipelines, primarily in repetitive genomic regions. This work introduces the utility of nanopore sequencing for MEI enrichment and lays the foundation for rapid discovery of elusive, repetitive genetic elements.

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Section: Cas9 Enrichment and Nanopore Sequencing Captures Non-referenmentioning

confidence: 87%

Cas9 targeted enrichment of mobile elements using nanopore sequencing

McDonald

Zhou

Castro

et al. 2021

Preprint

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“…For SNVs, we compared the calls from 3 strategies using the benchmark of NA12878 [40] and NA24385 [41]. For SVs, we compared 3 Linked-Read sets ( R 9 , R 10 , R 11 ) from HG002 with the Tier 1 SV benchmark from GIAB [42] and used VaPoR [43] to validate our SV calls based on PacBio CCS reads from NA24385 [44]. We compared SNV and SV calls among the different approaches using vcfeval [45] and truvari [42], respectively.…”

Section: Methodsmentioning

confidence: 99%

Assessment of human diploid genome assembly with 10x Linked-Reads data

et al. 2019

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Background Producing cost-effective haplotype-resolved personal genomes remains challenging. 10x Linked-Read sequencing, with its high base quality and long-range information, has been demonstrated to facilitate de novo assembly of human genomes and variant detection. In this study, we investigate in depth how the parameter space of 10x library preparation and sequencing affects assembly quality, on the basis of both simulated and real libraries. Results We prepared and sequenced eight 10x libraries with a diverse set of parameters from standard cell lines NA12878 and NA24385 and performed whole-genome assembly on the data. We also developed the simulator LRTK-SIM to follow the workflow of 10x data generation and produce realistic simulated Linked-Read data sets. We found that assembly quality could be improved by increasing the total sequencing coverage (C) and keeping physical coverage of DNA fragments (CF) or read coverage per fragment (CR) within broad ranges. The optimal physical coverage was between 332× and 823× and assembly quality worsened if it increased to >1,000× for a given C. Long DNA fragments could significantly extend phase blocks but decreased contig contiguity. The optimal length-weighted fragment length (W${\mu _{FL}}$) was ∼50–150 kb. When broadly optimal parameters were used for library preparation and sequencing, ∼80% of the genome was assembled in a diploid state. Conclusions The Linked-Read libraries we generated and the parameter space we identified provide theoretical considerations and practical guidelines for personal genome assemblies based on 10x Linked-Read sequencing.

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“…For computational validation, we obtained ONT reads of HG00733 from HGSVC and applied VaPoR [59], an independent structural variants validation method, to validate these CSVs ( Supplementary Note ). VaPoR is able to validate calls based predicted region and types with a confidence score.…”

Section: Methodsmentioning

confidence: 99%

Mako: a graph-based pattern growth approach to detect complex structural variants

Lin

Yang

Kosters

et al. 2021

Preprint

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Complex structural variants (CSVs) are genomic alterations that have more than two breakpoints and are considered as simultaneous occurrence of simple structural variants. However, detecting the compounded mutational signals of CSVs is challenging through a commonly used model-match strategy. As a result, there has been limited progress for CSV discovery compared with simple structural variants. We systematically analyzed the multi-breakpoint connection feature of CSVs, and proposed Mako, utilizing a bottom-up guided model-free strategy, to detect CSVs from paired-end short-read sequencing. Specifically, we implemented a graph-based pattern growth approach, where the graph depicts potential breakpoint connections and pattern growth enables CSV detection without predefined models. Comprehensive evaluations on both simulated and real datasets revealed that Mako outperformed other algorithms. Notably, validation rates of CSV on real data based on experimental and computational validations as well as manual inspections are around 70%, where the medians of experimental and computational breakpoint shift are 13bp and 26bp, respectively. Moreover, Mako CSV subgraph effectively characterized the breakpoint connections of a CSV event and uncovered a total of 15 CSV types, including two novel types of adjacent segments swap and tandem dispersed duplication. Further analysis of these CSVs also revealed impact of sequence homology in the formation of CSVs. Mako is publicly available at https://github.com/jiadong324/Mako.

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A recurrence based approach for validating structural variation using long-read sequencing technology

Cited by 13 publications

References 20 publications

Cas9 targeted enrichment of mobile elements using nanopore sequencing

Cas9 targeted enrichment of mobile elements using nanopore sequencing

Assessment of human diploid genome assembly with 10x Linked-Reads data

Mako: a graph-based pattern growth approach to detect complex structural variants

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