scholarly journals Limitations of lymphoblastoid cell lines for establishing genetic reference datasets in the immunoglobulin loci

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0261374
Author(s):  
Oscar L. Rodriguez ◽  
Andrew J. Sharp ◽  
Corey T. Watson
Keyword(s):  
Genetic Resources ◽  
Cell Lines ◽  
Association Studies ◽  
Population Level ◽  
Biomedical Science ◽  
Lymphoblastoid Cell Lines ◽  
Sequencing Data ◽  
Short Read ◽  
Short Read Sequencing ◽  
Genetics Research

Lymphoblastoid cell lines (LCLs) have been critical to establishing genetic resources for biomedical science. They have been used extensively to study human genetic diversity, genome function, and inform the development of tools and methodologies for augmenting disease genetics research. While the validity of variant callsets from LCLs has been demonstrated for most of the genome, previous work has shown that DNA extracted from LCLs is modified by V(D)J recombination within the immunoglobulin (IG) loci, regions that harbor antibody genes critical to immune system function. However, the impacts of V(D)J on short read sequencing data generated from LCLs has not been extensively investigated. In this study, we used LCL-derived short read sequencing data from the 1000 Genomes Project (n = 2,504) to identify signatures of V(D)J recombination. Our analyses revealed sample-level impacts of V(D)J recombination that varied depending on the degree of inferred monoclonality. We showed that V(D)J associated somatic deletions impacted genotyping accuracy, leading to adulterated population-level estimates of allele frequency and linkage disequilibrium. These findings illuminate limitations of using LCLs and short read data for building genetic resources in the IG loci, with implications for interpreting previous disease association studies in these regions.

scholarly journals Limitations of lymphoblastoid cell lines for establishing genetic reference datasets in the immunoglobulin loci

2021 ◽  
Author(s):  
Oscar L Rodriguez ◽  
Andrew J Sharp ◽  
Corey T Watson
Keyword(s):  
Genetic Resources ◽  
Cell Lines ◽  
Association Studies ◽  
Population Level ◽  
Biomedical Science ◽  
Lymphoblastoid Cell Lines ◽  
Sequencing Data ◽  
Short Read Sequencing ◽  
Genetics Research ◽  
Previous Disease

Lymphoblastoid cell lines (LCLs) have been critical to establishing genetic resources for biomedical science. They have been used extensively to study human genetic diversity, genome function, and inform the development of tools and methodologies for augmenting disease genetics research. While the validity of variant callsets from LCLs has been demonstrated for most of the genome, previous work has shown that DNA extracted from LCLs is modified by V(D)J recombination within the immunoglobulin (IG) loci, regions that harbor antibody genes critical to immune system function. However, the impacts of V(D)J on data generated from LCLs has not been extensively investigated. In this study, we used LCL-derived short read sequencing data from the 1000 Genomes Project (n=2,504) to identify signatures of V(D)J recombination. Our analyses revealed sample-level impacts of V(D)J recombination that varied depending on the degree of inferred monoclonality. We showed that V(D)J associated somatic deletions impacted genotyping accuracy, leading to adulterated population-level estimates of allele frequency and linkage disequilibrium. These findings illuminate limitations of using LCLs for building genetic resources in the IG loci, with implications for interpreting previous disease association studies in these regions.

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

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).

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

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.

scholarly journals Rapid Mycobacterium tuberculosis spoligotyping from uncorrected long reads using Galru

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.

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

Cells ◽  
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.

scholarly journals Complete Genome Sequence of Rubrobacter xylanophilus Strain AA3-22, Isolated from Arima Onsen in Japan

2019 ◽  
Vol 8 (34) ◽  
Author(s):  
Natsuki Tomariguchi ◽  
Kentaro Miyazaki
Keyword(s):  
Genome Sequence ◽  
Complete Genome Sequence ◽  
Complete Genome ◽  
Hot Spring ◽  
Sequencing Data ◽  
Short Read ◽  
Content Type ◽  
Short Read Sequencing ◽  
Oxford Nanopore ◽  
Long Read

Rubrobacter xylanophilus strain AA3-22, belonging to the phylum Actinobacteria, was isolated from nonvolcanic Arima Onsen (hot spring) in Japan. Here, we report the complete genome sequence of this organism, which was obtained by combining Oxford Nanopore long-read and Illumina short-read sequencing data.

scholarly journals Fast and accurate HLA typing from short-read next-generation sequence data with xHLA

2017 ◽  
Vol 114 (30) ◽  
pp. 8059-8064 ◽  
Author(s):  
Chao Xie ◽  
Zhen Xuan Yeo ◽  
Marie Wong ◽  
Jason Piper ◽  
Tao Long ◽  
...  
Keyword(s):  
Sequence Data ◽  
Sequence Similarity ◽  
Amino Acid Level ◽  
Hla Typing ◽  
Sequencing Data ◽  
Desktop Computer ◽  
Short Read ◽  
Short Read Sequencing ◽  
Hla Genes ◽  
Human Chromosome 6

The HLA gene complex on human chromosome 6 is one of the most polymorphic regions in the human genome and contributes in large part to the diversity of the immune system. Accurate typing of HLA genes with short-read sequencing data has historically been difficult due to the sequence similarity between the polymorphic alleles. Here, we introduce an algorithm, xHLA, that iteratively refines the mapping results at the amino acid level to achieve 99–100% four-digit typing accuracy for both class I and II HLA genes, taking only∼3 min to process a 30× whole-genome BAM file on a desktop computer.

scholarly journals Indel variant analysis of short-read sequencing data with Scalpel

2016 ◽  
Vol 11 (12) ◽  
pp. 2529-2548 ◽  
Author(s):  
Han Fang ◽  
Ewa A Bergmann ◽  
Kanika Arora ◽  
Vladimir Vacic ◽  
Michael C Zody ◽  
...  
Keyword(s):  
Sequencing Data ◽  
Short Read ◽  
Short Read Sequencing ◽  
Variant Analysis

scholarly journals LinkedSV for detection of mosaic structural variants from linked-read exome and genome sequencing data

2018 ◽  
Author(s):  
Li Fang ◽  
Charlly Kao ◽  
Michael V Gonzalez ◽  
Fernanda A Mafra ◽  
Renata Pellegrino da Silva ◽  
...  
Keyword(s):  
Exome Sequencing ◽  
Read Depth ◽  
Structural Variants ◽  
Sequencing Data ◽  
High Coverage ◽  
Short Read ◽  
Short Read Sequencing ◽  
Sequencing Studies ◽  
Long Read ◽  
Local Assembly

AbstractLinked-read sequencing provides long-range information on short-read sequencing data by barcoding reads originating from the same DNA molecule, and can improve the detection and breakpoint identification for structural variants (SVs). We present LinkedSV for SV detection on linked-read sequencing data. LinkedSV considers barcode overlapping and enriched fragment endpoints as signals to detect large SVs, while it leverages read depth, paired-end signals and local assembly to detect small SVs. Benchmarking studies demonstrates that LinkedSV outperforms existing tools, especially on exome data and on somatic SVs with low variant allele frequencies. We demonstrate clinical cases where LinkedSV identifies disease causal SVs from linked-read exome sequencing data missed by conventional exome sequencing, and show examples where LinkedSV identifies SVs missed by high-coverage long-read sequencing. In summary, LinkedSV can detect SVs missed by conventional short-read and long-read sequencing approaches, and may resolve negative cases from clinical genome/exome sequencing studies.

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