Long-read sequencing identified a novel nonsense and a de novo missense of PPA2 in trans in a Chinese patient with autosomal recessive infantile sudden cardiac failure

Lipoid congenital adrenal hyperplasia (LCAH) is typically inherited as an autosomal recessive condition. There are 3 reports of individuals with a dominantly acting heterozygous variant leading to a clinically significant phenotype. We report a 46,XY child with a novel heterozygous intronic variant in STAR resulting in LCAH with an attenuated genital phenotype. The patient presented with neonatal hypoglycemia and had descended testes with a fused scrotum and small phallus. Evaluation revealed primary adrenal insufficiency with deficiencies of cortisol, aldosterone, and androgens. He was found to have a de novo heterozygous novel variant in STAR: c.65-2A>C. We report a case of a novel variant and review of other dominant mutations at the same position in the literature. Clinicians should be aware of the possibility of attenuated genital phenotypes of LCAH and the contribution of de novo variants in STAR at c.65-2 to the pathogenesis of that phenotype.

Download Full-text

Long‐read sequencing and de novo assembly of the Luffa cylindrica (L.) Roem. genome

Molecular Ecology Resources ◽

10.1111/1755-0998.13129 ◽

2020 ◽

Vol 20 (2) ◽

pp. 511-519 ◽

Cited By ~ 5

Author(s):

Tao Zhang ◽

Xuyan Ren ◽

Zhao Zhang ◽

Yao Ming ◽

Zhe Yang ◽

...

Keyword(s):

De Novo Assembly ◽

De Novo ◽

Luffa Cylindrica ◽

Long Read

Download Full-text

Genotyping structural variants in pangenome graphs using the vg toolkit

10.1101/654566 ◽

2019 ◽

Cited By ~ 7

Author(s):

Glenn Hickey ◽

David Heller ◽

Jean Monlong ◽

Jonas A. Sibbesen ◽

Jouni Sirén ◽

...

Keyword(s):

De Novo ◽

State Of The Art ◽

Effective Means ◽

Point Mutations ◽

Structural Variants ◽

Short Read ◽

Yeast Strains ◽

Sequencing Studies ◽

Long Read

AbstractStructural variants (SVs) remain challenging to represent and study relative to point mutations despite their demonstrated importance. We show that variation graphs, as implemented in the vg toolkit, provide an effective means for leveraging SV catalogs for short-read SV genotyping experiments. We benchmarked vg against state-of-the-art SV genotypers using three sequence-resolved SV catalogs generated by recent long-read sequencing studies. In addition, we use assemblies from 12 yeast strains to show that graphs constructed directly from aligned de novo assemblies improve genotyping compared to graphs built from intermediate SV catalogs in the VCF format.

Download Full-text

Chromosome-level assembly of Drosophila bifasciata reveals important karyotypic transition of the X chromosome

10.1101/847558 ◽

2019 ◽

Author(s):

Ryan Bracewell ◽

Anita Tran ◽

Kamalakar Chatla ◽

Doris Bachtrog

Keyword(s):

X Chromosome ◽

Genome Assembly ◽

De Novo ◽

Pericentromeric Region ◽

Species Group ◽

Chromosome 15 ◽

Protein Coding ◽

Protein Coding Genes ◽

Long Read ◽

Chromosome Level

ABSTRACTThe Drosophila obscura species group is one of the most studied clades of Drosophila and harbors multiple distinct karyotypes. Here we present a de novo genome assembly and annotation of D. bifasciata, a species which represents an important subgroup for which no high-quality chromosome-level genome assembly currently exists. We combined long-read sequencing (Nanopore) and Hi-C scaffolding to achieve a highly contiguous genome assembly approximately 193Mb in size, with repetitive elements constituting 30.1% of the total length. Drosophila bifasciata harbors four large metacentric chromosomes and the small dot, and our assembly contains each chromosome in a single scaffold, including the highly repetitive pericentromere, which were largely composed of Jockey and Gypsy transposable elements. We annotated a total of 12,821 protein-coding genes and comparisons of synteny with D. athabasca orthologs show that the large metacentric pericentromeric regions of multiple chromosomes are conserved between these species. Importantly, Muller A (X chromosome) was found to be metacentric in D. bifasciata and the pericentromeric region appears homologous to the pericentromeric region of the fused Muller A-AD (XL and XR) of pseudoobscura/affinis subgroup species. Our finding suggests a metacentric ancestral X fused to a telocentric Muller D and created the large neo-X (Muller A-AD) chromosome ∼15 MYA. We also confirm the fusion of Muller C and D in D. bifasciata and show that it likely involved a centromere-centromere fusion.

Download Full-text

Accurate long-read de novo assembly evaluation with Inspector

Genome Biology ◽

10.1186/s13059-021-02527-4 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Yu Chen ◽

Yixin Zhang ◽

Amy Y. Wang ◽

Min Gao ◽

Zechen Chong

Keyword(s):

Genome Assembly ◽

De Novo Assembly ◽

In Silico ◽

Large Scale ◽

De Novo ◽

Small Scale ◽

De Novo Genome Assembly ◽

Consensus Sequences ◽

Assembly Evaluation ◽

Long Read

AbstractLong-read de novo genome assembly continues to advance rapidly. However, there is a lack of effective tools to accurately evaluate the assembly results, especially for structural errors. We present Inspector, a reference-free long-read de novo assembly evaluator which faithfully reports types of errors and their precise locations. Notably, Inspector can correct the assembly errors based on consensus sequences derived from raw reads covering erroneous regions. Based on in silico and long-read assembly results from multiple long-read data and assemblers, we demonstrate that in addition to providing generic metrics, Inspector can accurately identify both large-scale and small-scale assembly errors.

Download Full-text

Contiguity: Contig adjacency graph construction and visualisation

10.7287/peerj.preprints.1037v1 ◽

2015 ◽

Cited By ~ 8

Author(s):

Mitchell J Sullivan ◽

Nouri L Ben Zakour ◽

Brian M Forde ◽

Mitchell Stanton-Cook ◽

Scott A Beatson

Keyword(s):

De Novo ◽

Reference Sequence ◽

De Bruijn Graph ◽

Interactive Software ◽

Graph Exploration ◽

Adjacency Graph ◽

Highly Sensitive ◽

Long Read ◽

Genome Assemblies ◽

Adjacency Graphs

Contiguity is an interactive software for the visualization and manipulation of de novo genome assemblies. Contiguity creates and displays information on contig adjacency which is contextualized by the simultaneous display of a comparison between assembled contigs and reference sequence. Where scaffolders allow unambiguous connections between contigs to be resolved into a single scaffold, Contiguity allows the user to create all potential scaffolds in ambiguous regions of the genome. This enables the resolution of novel sequence or structural variants from the assembly. In addition, Contiguity provides a sequencing and assembly agnostic approach for the creation of contig adjacency graphs. To maximize the number of contig adjacencies determined, Contiguity combines information from read pair mappings, sequence overlap and De Bruijn graph exploration. We demonstrate how highly sensitive graphs can be achieved using this method. Contig adjacency graphs allow the user to visualize potential arrangements of contigs in unresolvable areas of the genome. By combining adjacency information with comparative genomics, Contiguity provides an intuitive approach for exploring and improving sequence assemblies. It is also useful in guiding manual closure of long read sequence assemblies. Contiguity is an open source application, implemented using Python and the Tkinter GUI package that can run on any Unix, OSX and Windows operating system. It has been designed and optimized for bacterial assemblies. Contiguity is available at http://mjsull.github.io/Contiguity .

Download Full-text

LRSim: a Linked Reads Simulator generating insights for better genome partitioning

10.1101/103549 ◽

2017 ◽

Cited By ~ 1

Author(s):

Ruibang Luo ◽

Fritz J. Sedlazeck ◽

Charlotte A. Darby ◽

Stephen M. Kelly ◽

Michael C. Schatz

Keyword(s):

Genome Assembly ◽

Fragment Length ◽

De Novo ◽

Library Preparation ◽

Haplotype Phasing ◽

Multiple Datasets ◽

Long Read ◽

Fine Control ◽

Informatics Tools ◽

Sequencing Process

AbstractMotivationLinked reads are a form of DNA sequencing commercialized by 10X Genomics that uses highly multiplexed barcoding within microdroplets to tag short reads to progenitor molecules. The linked reads, spanning tens to hundreds of kilobases, offer an alternative to long-read sequencing for de novo assembly, haplotype phasing and other applications. However, there is no available simulator, making it difficult to measure their capability or develop new informatics tools.ResultsOur analysis of 13 real linked read datasets revealed their characteristics of barcodes, molecules and partitions. Based on this, we introduce LRSim that simulates linked reads by emulating the library preparation and sequencing process with fine control of 1) the number of simulated variants; 2) the linked-read characteristics; and 3) the Illumina reads profile. We conclude from the phasing and genome assembly of multiple datasets, recommendations on coverage, fragment length, and partitioning when sequencing human and non-human genome.AvailabilityLRSIM is under MIT license and is freely available at https://github.com/aquaskyline/[email protected]

Download Full-text

Mapping and phasing of structural variation in patient genomes using nanopore sequencing

10.1101/129379 ◽

2017 ◽

Cited By ~ 4

Author(s):

Mircea Cretu Stancu ◽

Markus J. van Roosmalen ◽

Ivo Renkens ◽

Marleen Nieboer ◽

Sjors Middelkamp ◽

...

Keyword(s):

Single Molecule ◽

De Novo ◽

Structural Variants ◽

Human Genetic Disease ◽

Structural Genomic ◽

Short Read ◽

Sequencing Technologies ◽

Genome Wide ◽

Long Read ◽

Complex Structural

AbstractStructural genomic variants form a common type of genetic alteration underlying human genetic disease and phenotypic variation. Despite major improvements in genome sequencing technology and data analysis, the detection of structural variants still poses challenges, particularly when variants are of high complexity. Emerging long-read single-molecule sequencing technologies provide new opportunities for detection of structural variants. Here, we demonstrate sequencing of the genomes of two patients with congenital abnormalities using the ONT MinION at 11x and 16x mean coverage, respectively. We developed a bioinformatic pipeline - NanoSV - to efficiently map genomic structural variants (SVs) from the long-read data. We demonstrate that the nanopore data are superior to corresponding short-read data with regard to detection of de novo rearrangements originating from complex chromothripsis events in the patients. Additionally, genome-wide surveillance of SVs, revealed 3,253 (33%) novel variants that were missed in short-read data of the same sample, the majority of which are duplications < 200bp in size. Long sequencing reads enabled efficient phasing of genetic variations, allowing the construction of genome-wide maps of phased SVs and SNVs. We employed read-based phasing to show that all de novo chromothripsis breakpoints occurred on paternal chromosomes and we resolved the long-range structure of the chromothripsis. This work demonstrates the value of long-read sequencing for screening whole genomes of patients for complex structural variants.

Download Full-text

Oxford Nanopore Sequencing, Hybrid Error Correction, and de novo Assembly of a Eukaryotic Genome

10.1101/013490 ◽

2015 ◽

Cited By ~ 23

Author(s):

Sara Goodwin ◽

James Gurtowski ◽

Scott Ethe-Sayers ◽

Panchajanya Deshpande ◽

Michael Schatz ◽

...

Keyword(s):

Error Correction ◽

De Novo Assembly ◽

De Novo ◽

Correction Algorithm ◽

Membrane Pore ◽

Complete Representation ◽

Oxford Nanopore ◽

Long Read ◽

Error Correction Algorithm ◽

Sequencing Instrument

Monitoring the progress of DNA molecules through a membrane pore has been postulated as a method for sequencing DNA for several decades. Recently, a nanopore-based sequencing instrument, the Oxford Nanopore MinION, has become available that we used for sequencing the S. cerevisiae genome. To make use of these data, we developed a novel open-source hybrid error correction algorithm Nanocorr (https://github.com/jgurtowski/nanocorr) specifically for Oxford Nanopore reads, as existing packages were incapable of assembling the long read lengths (5-50kbp) at such high error rate (between ~5 and 40% error). With this new method we were able to perform a hybrid error correction of the nanopore reads using complementary MiSeq data and produce a de novo assembly that is highly contiguous and accurate: the contig N50 length is more than ten-times greater than an Illumina-only assembly (678kb versus 59.9kbp), and has greater than 99.88% consensus identity when compared to the reference. Furthermore, the assembly with the long nanopore reads presents a much more complete representation of the features of the genome and correctly assembles gene cassettes, rRNAs, transposable elements, and other genomic features that were almost entirely absent in the Illumina-only assembly.

Download Full-text

De novo assembly of a Tibetan genome and identification of novel structural variants associated with high-altitude adaptation

National Science Review ◽

10.1093/nsr/nwz160 ◽

2019 ◽

Vol 7 (2) ◽

pp. 391-402 ◽

Cited By ~ 3

Author(s):

Yaoxi He ◽

Haiyi Lou ◽

Chaoying Cui ◽

Lian Deng ◽

Yang Gao ◽

...

Keyword(s):

High Altitude ◽

De Novo Assembly ◽

De Novo ◽

Population Analysis ◽

Extreme Environments ◽

Structural Variants ◽

Base Pairs ◽

High Quality ◽

Long Read ◽

Altitude Adaptation

Abstract Structural variants (SVs) may play important roles in human adaptation to extreme environments such as high altitude but have been under-investigated. Here, combining long-read sequencing with multiple scaffolding techniques, we assembled a high-quality Tibetan genome (ZF1), with a contig N50 length of 24.57 mega-base pairs (Mb) and a scaffold N50 length of 58.80 Mb. The ZF1 assembly filled 80 remaining N-gaps (0.25 Mb in total length) in the reference human genome (GRCh38). Markedly, we detected 17 900 SVs, among which the ZF1-specific SVs are enriched in GTPase activity that is required for activation of the hypoxic pathway. Further population analysis uncovered a 163-bp intronic deletion in the MKL1 gene showing large divergence between highland Tibetans and lowland Han Chinese. This deletion is significantly associated with lower systolic pulmonary arterial pressure, one of the key adaptive physiological traits in Tibetans. Moreover, with the use of the high-quality de novo assembly, we observed a much higher rate of genome-wide archaic hominid (Altai Neanderthal and Denisovan) shared non-reference sequences in ZF1 (1.32%–1.53%) compared to other East Asian genomes (0.70%–0.98%), reflecting a unique genomic composition of Tibetans. One such archaic hominid shared sequence—a 662-bp intronic insertion in the SCUBE2 gene—is enriched and associated with better lung function (the FEV1/FVC ratio) in Tibetans. Collectively, we generated the first high-resolution Tibetan reference genome, and the identified SVs may serve as valuable resources for future evolutionary and medical studies.

Download Full-text