ExpansionHunter Denovo: A computational method for locating known and novel repeat expansions in short-read sequencing data

AbstractExpansions of short tandem repeats are responsible for over 40 monogenic disorders, and undoubtedly many more pathogenic repeat expansions (REs) remain to be discovered. Existing methods for detecting REs in short-read sequencing data require predefined repeat catalogs. However recent discoveries have emphasized the need for detection methods that do not require candidate repeats to be specified in advance. To address this need, we introduce ExpansionHunter Denovo, an efficient catalog-free method for genome-wide detection of REs. Analysis of real and simulated data shows that our method can identify large expansions of 41 out of 44 pathogenic repeats, including nine recently reported non-reference REs not discoverable via existing methods.ExpansionHunter Denovo is freely available at https://github.com/Illumina/ExpansionHunterDenovo

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ExpansionHunter Denovo: a computational method for locating known and novel repeat expansions in short-read sequencing data

Genome Biology ◽

10.1186/s13059-020-02017-z ◽

2020 ◽

Vol 21 (1) ◽

Cited By ~ 6

Author(s):

Egor Dolzhenko ◽

Mark F. Bennett ◽

Phillip A. Richmond ◽

Brett Trost ◽

Sai Chen ◽

...

Keyword(s):

Computational Method ◽

Sequencing Data ◽

Short Read ◽

Short Read Sequencing ◽

Repeat Expansions

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REscan: inferring repeat expansions and structural variation in paired-end short read sequencing data

Bioinformatics ◽

10.1093/bioinformatics/btaa753 ◽

2020 ◽

Author(s):

Russell Lewis McLaughlin

Keyword(s):

Structural Variation ◽

Sequence Data ◽

Neurological Diseases ◽

Repeat Expansion ◽

Sequencing Data ◽

Short Read ◽

Short Read Sequencing ◽

Repeat Expansions ◽

Paired End Sequencing

Abstract Motivation Repeat expansions are an important class of genetic variation in neurological diseases. However, the identification of novel repeat expansions using conventional sequencing methods is a challenge due to their typical lengths relative to short sequence reads and difficulty in producing accurate and unique alignments for repetitive sequence. However, this latter property can be harnessed in paired-end sequencing data to infer the possible locations of repeat expansions and other structural variation. Results This article presents REscan, a command-line utility that infers repeat expansion loci from paired-end short read sequencing data by reporting the proportion of reads orientated towards a locus that do not have an adequately mapped mate. A high REscan statistic relative to a population of data suggests a repeat expansion locus for experimental follow-up. This approach is validated using genome sequence data for 259 cases of amyotrophic lateral sclerosis, of which 24 are positive for a large repeat expansion in C9orf72, showing that REscan statistics readily discriminate repeat expansion carriers from non-carriers. Availabilityand implementation C source code at https://github.com/rlmcl/rescan (GNU General Public Licence v3).

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Critical assessment of bioinformatics methods for the characterization of pathological repeat expansions with single-molecule sequencing data

Briefings in Bioinformatics ◽

10.1093/bib/bbz099 ◽

2019 ◽

Vol 21 (6) ◽

pp. 1971-1986 ◽

Cited By ~ 1

Author(s):

Matteo Chiara ◽

Federico Zambelli ◽

Ernesto Picardi ◽

David S Horner ◽

Graziano Pesole

Keyword(s):

Single Molecule ◽

Tandem Repeats ◽

Simulated Data ◽

Detailed Comparison ◽

Sequencing Data ◽

Single Molecule Sequencing ◽

Sequencing Technologies ◽

Repeat Expansions

Abstract A number of studies have reported the successful application of single-molecule sequencing technologies to the determination of the size and sequence of pathological expanded microsatellite repeats over the last 5 years. However, different custom bioinformatics pipelines were employed in each study, preventing meaningful comparisons and somewhat limiting the reproducibility of the results. In this review, we provide a brief summary of state-of-the-art methods for the characterization of expanded repeats alleles, along with a detailed comparison of bioinformatics tools for the determination of repeat length and sequence, using both real and simulated data. Our reanalysis of publicly available human genome sequencing data suggests a modest, but statistically significant, increase of the error rate of single-molecule sequencing technologies at genomic regions containing short tandem repeats. However, we observe that all the methods herein tested, irrespective of the strategy used for the analysis of the data (either based on the alignment or assembly of the reads), show high levels of sensitivity in both the detection of expanded tandem repeats and the estimation of the expansion size, suggesting that approaches based on single-molecule sequencing technologies are highly effective for the detection and quantification of tandem repeat expansions and contractions.

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REViewer: Haplotype-resolved visualization of read alignments in and around tandem repeats

10.1101/2021.10.20.465046 ◽

2021 ◽

Author(s):

Egor Dolzhenko ◽

Ben Weisburd ◽

Kristina Ibanez ◽

Indhu Shree Rajan Babu ◽

Mark F Bennett ◽

...

Keyword(s):

Tandem Repeats ◽

Visual Assessment ◽

Repeat Sequence ◽

Computational Method ◽

Triplet Repeat ◽

Sequencing Data ◽

Training Materials ◽

Repeat Expansions ◽

Genomic Regions ◽

Flanking Regions

Background: Expansions of short tandem repeats are the cause of many neurogenetic disorders including familial amyotrophic lateral sclerosis, Huntington disease, and many others. Multiple methods have been recently developed that can identify repeat expansions in whole genome or exome sequencing data. Despite the widely-recognized need for visual assessment of variant calls in clinical settings, current computational tools lack the ability to produce such visualizations for repeat expansions. Expanded repeats are difficult to visualize because they correspond to large insertions relative to the reference genome and involve many misaligning and ambiguously aligning reads. Results: We implemented REViewer, a computational method for visualization of sequencing data in genomic regions containing long repeat expansions. To generate a read pileup, REViewer reconstructs local haplotype sequences and distributes reads to these haplotypes in a way that is most consistent with the fragment lengths and evenness of read coverage. To create appropriate training materials for onboarding new users, we performed a concordance study involving 12 scientists involved in STR research. We used the results of this study to create a user guide that describes the basic principles of using REViewer as well as a guide to the typical features of read pileups that correspond to low confidence repeat genotype calls. Additionally, we demonstrated that REViewer can be used to annotate clinically-relevant repeat interruptions by comparing visual assessment results of 44 FMR1 repeat alleles with the results of triplet repeat primed PCR. For 38 of these alleles, the results of visual assessment were consistent with triplet repeat primed PCR. Conclusions: Read pileup plots generated by REViewer offer an intuitive way to visualize sequencing data in regions containing long repeat expansions. Laboratories can use REViewer to assess the quality of repeat genotype calls as well as to visually detect interruptions or other imperfections in the repeat sequence and the surrounding flanking regions.

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Detecting Expansions of Tandem Repeats in Cohorts Sequenced with Short-Read Sequencing Data

The American Journal of Human Genetics ◽

10.1016/j.ajhg.2018.10.015 ◽

2018 ◽

Vol 103 (6) ◽

pp. 858-873 ◽

Cited By ~ 34

Author(s):

Rick M. Tankard ◽

Mark F. Bennett ◽

Peter Degorski ◽

Martin B. Delatycki ◽

Paul J. Lockhart ◽

...

Keyword(s):

Tandem Repeats ◽

Sequencing Data ◽

Short Read ◽

Short Read Sequencing

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High resolution copy number inference in cancer using short-molecule nanopore sequencing

10.1101/2020.12.28.424602 ◽

2020 ◽

Author(s):

Timour Baslan ◽

Sam Kovaka ◽

Fritz J. Sedlazeck ◽

Yanming Zhang ◽

Robert Wappel ◽

...

Keyword(s):

Copy Number ◽

Cost Effective ◽

Chromosome Analysis ◽

Ease Of Use ◽

Precision Oncology ◽

Nanopore Sequencing ◽

Dna Molecules ◽

Sequencing Data ◽

Short Read ◽

Short Read Sequencing

ABSTRACTGenome copy number is an important source of genetic variation in health and disease. In cancer, clinically actionable Copy Number Alterations (CNAs) can be inferred from short-read sequencing data, enabling genomics-based precision oncology. Emerging Nanopore sequencing technologies offer the potential for broader clinical utility, for example in smaller hospitals, due to lower instrument cost, higher portability, and ease of use. Nonetheless, Nanopore sequencing devices are limited in terms of the number of retrievable sequencing reads/molecules compared to short-read sequencing platforms. This represents a challenge for applications that require high read counts such as CNA inference. To address this limitation, we targeted the sequencing of short-length DNA molecules loaded at optimized concentration in an effort to increase sequence read/molecule yield from a single nanopore run. We show that sequencing short DNA molecules reproducibly returns high read counts and allows high quality CNA inference. We demonstrate the clinical relevance of this approach by accurately inferring CNAs in acute myeloid leukemia samples. The data shows that, compared to traditional approaches such as chromosome analysis/cytogenetics, short molecule nanopore sequencing returns more sensitive, accurate copy number information in a cost effective and expeditious manner, including for multiplex samples. Our results provide a framework for the sequencing of relatively short DNA molecules on nanopore devices with applications in research and medicine, that include but are not limited to, CNAs.

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EagleImp: Fast and Accurate Genome-wide Phasing and Imputation in a Single Tool

10.1101/2022.01.11.475810 ◽

2022 ◽

Author(s):

Lars Wienbrandt ◽

David Ellinghaus

Keyword(s):

Memory Management ◽

Imputation Accuracy ◽

Simulated Data ◽

Genotype Imputation ◽

Whole Genome Sequencing Data ◽

Common Variants ◽

Sequencing Data ◽

1000 Genomes ◽

Genome Wide ◽

Reference Genomes

Background: Reference-based phasing and genotype imputation algorithms have been developed with sublinear theoretical runtime behaviour, but runtimes are still high in practice when large genome-wide reference datasets are used. Methods: We developed EagleImp, a software with algorithmic and technical improvements and new features for accurate and accelerated phasing and imputation in a single tool. Results: We compared accuracy and runtime of EagleImp with Eagle2, PBWT and prominent imputation servers using whole-genome sequencing data from the 1000 Genomes Project, the Haplotype Reference Consortium and simulated data with more than 1 million reference genomes. EagleImp is 2 to 10 times faster (depending on the single or multiprocessor configuration selected) than Eagle2/PBWT, with the same or better phasing and imputation quality in all tested scenarios. For common variants investigated in typical GWAS studies, EagleImp provides same or higher imputation accuracy than the Sanger Imputation Service, Michigan Imputation Server and the newly developed TOPMed Imputation Server, despite larger (not publicly available) reference panels. It has many new features, including automated chromosome splitting and memory management at runtime to avoid job aborts, fast reading and writing of large files, and various user-configurable algorithm and output options. Conclusions: Due to the technical optimisations, EagleImp can perform fast and accurate reference-based phasing and imputation for future very large reference panels with more than 1 million genomes. EagleImp is freely available for download from https://github.com/ikmb/eagleimp.

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Rapid Mycobacterium tuberculosis spoligotyping from uncorrected long reads using Galru

10.1101/2020.05.31.126490 ◽

2020 ◽

Author(s):

Andrew J. Page ◽

Nabil-Fareed Alikhan ◽

Michael Strinden ◽

Thanh Le Viet ◽

Timofey Skvortsov

Keyword(s):

Mycobacterium Tuberculosis ◽

State Of The Art ◽

Sequence Data ◽

Human Pathogen ◽

Sequencing Data ◽

Short Read ◽

Short Read Sequencing ◽

Long Reads ◽

Long Read

AbstractSpoligotyping of Mycobacterium tuberculosis provides a subspecies classification of this major human pathogen. Spoligotypes can be predicted from short read genome sequencing data; however, no methods exist for long read sequence data such as from Nanopore or PacBio. We present a novel software package Galru, which can rapidly detect the spoligotype of a Mycobacterium tuberculosis sample from as little as a single uncorrected long read. It allows for near real-time spoligotyping from long read data as it is being sequenced, giving rapid sample typing. We compare it to the existing state of the art software and find it performs identically to the results obtained from short read sequencing data. Galru is freely available from https://github.com/quadram-institute-bioscience/galru under the GPLv3 open source licence.

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Transposable element expression in tumors is associated with immune infiltration and increased antigenicity

Nature Communications ◽

10.1038/s41467-019-13035-2 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 16

Author(s):

Yu Kong ◽

Christopher M. Rose ◽

Ashley A. Cass ◽

Alexander G. Williams ◽

Martine Darwish ◽

...

Keyword(s):

Dna Methylation ◽

De Novo ◽

Computational Method ◽

The Cancer Genome Atlas ◽

Potential Consequence ◽

Sequencing Data ◽

Antiviral Responses ◽

Genome Wide ◽

Cancer Genome Atlas ◽

Demethylation Agent

AbstractProfound global loss of DNA methylation is a hallmark of many cancers. One potential consequence of this is the reactivation of transposable elements (TEs) which could stimulate the immune system via cell-intrinsic antiviral responses. Here, we develop REdiscoverTE, a computational method for quantifying genome-wide TE expression in RNA sequencing data. Using The Cancer Genome Atlas database, we observe increased expression of over 400 TE subfamilies, of which 262 appear to result from a proximal loss of DNA methylation. The most recurrent TEs are among the evolutionarily youngest in the genome, predominantly expressed from intergenic loci, and associated with antiviral or DNA damage responses. Treatment of glioblastoma cells with a demethylation agent results in both increased TE expression and de novo presentation of TE-derived peptides on MHC class I molecules. Therapeutic reactivation of tumor-specific TEs may synergize with immunotherapy by inducing inflammation and the display of potentially immunogenic neoantigens.

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A Transposon Story: From TE Content to TE Dynamic Invasion of Drosophila Genomes Using the Single-Molecule Sequencing Technology from Oxford Nanopore

Cells ◽

10.3390/cells9081776 ◽

2020 ◽

Vol 9 (8) ◽

pp. 1776

Author(s):

Mourdas Mohamed ◽

Nguyet Thi-Minh Dang ◽

Yuki Ogyama ◽

Nelly Burlet ◽

Bruno Mugat ◽

...

Keyword(s):

Single Molecule ◽

Wild Type ◽

Sequencing Data ◽

Short Read ◽

Short Read Sequencing ◽

Sequencing Technologies ◽

Oxford Nanopore ◽

In The Wild ◽

Successive Generations ◽

Type Strains

Transposable elements (TEs) are the main components of genomes. However, due to their repetitive nature, they are very difficult to study using data obtained with short-read sequencing technologies. Here, we describe an efficient pipeline to accurately recover TE insertion (TEI) sites and sequences from long reads obtained by Oxford Nanopore Technology (ONT) sequencing. With this pipeline, we could precisely describe the landscapes of the most recent TEIs in wild-type strains of Drosophila melanogaster and Drosophila simulans. Their comparison suggests that this subset of TE sequences is more similar than previously thought in these two species. The chromosome assemblies obtained using this pipeline also allowed recovering piRNA cluster sequences, which was impossible using short-read sequencing. Finally, we used our pipeline to analyze ONT sequencing data from a D. melanogaster unstable line in which LTR transposition was derepressed for 73 successive generations. We could rely on single reads to identify new insertions with intact target site duplications. Moreover, the detailed analysis of TEIs in the wild-type strains and the unstable line did not support the trap model claiming that piRNA clusters are hotspots of TE insertions.

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