Draft genome assemblies using sequencing reads from Oxford Nanopore Technology and Illumina platforms for four species of North American Fundulus killifish

Abstract Background Whole-genome sequencing data from wild-caught individuals of closely related North American killifish species (Fundulus xenicus, Fundulus catenatus, Fundulus nottii, and Fundulus olivaceus) were obtained using long-read Oxford Nanopore Technology (ONT) PromethION and short-read Illumina platforms. Findings Draft de novo reference genome assemblies were generated using a combination of long and short sequencing reads. For each species, the PromethION platform was used to generate 30–45× sequence coverage, and the Illumina platform was used to generate 50–160× sequence coverage. Illumina-only assemblies were fragmented with high numbers of contigs, while ONT-only assemblies were error prone with low BUSCO scores. The highest N50 values, ranging from 0.4 to 2.7 Mb, were from assemblies generated using a combination of short- and long-read data. BUSCO scores were consistently >90% complete using the Eukaryota database. Conclusions High-quality genomes can be obtained from a combination of using short-read Illumina data to polish assemblies generated with long-read ONT data. Draft assemblies and raw sequencing data are available for public use. We encourage use and reuse of these data for assembly benchmarking and other analyses.

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Draft genome assemblies using sequencing reads from Oxford Nanopore Technology and Illumina platforms for four species of North American killifish from the Fundulus genus

10.1101/686246 ◽

2019 ◽

Author(s):

Lisa K. Johnson ◽

Ruta Sahasrabudhe ◽

Tony Gill ◽

Jennifer Roach ◽

Lutz Froenicke ◽

...

Keyword(s):

North American ◽

De Novo ◽

Draft Genome ◽

Sequencing Data ◽

Sequence Coverage ◽

Illumina Platform ◽

Oxford Nanopore ◽

Fundulus Olivaceus ◽

Genome Assemblies ◽

Oxford Nanopore Technologies

AbstractDraft de novo reference genome assemblies were obtained from four North American killifish species (Fundulus xenicus, Fundulus catenatus, Fundulus nottii, and Fundulus olivaceus) using sequence reads from Illumina and Oxford Nanopore Technologies’ PromethION platforms. For each species, the PromethION platform was used to generate 30-45x sequence coverage, and the Illumina platform was used to generate 50-160x sequence coverage. Contig N50 values ranged from 0.4 Mb to 2.7 Mb, and BUSCO scores were consistently above 90% complete using the Eukaryota database. Draft assemblies and raw sequencing data are available for public use. We encourage use and re-use of these data for assembly benchmarking and external analyses.

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Comparison of long read methods for sequencing and assembly of a plant genome

10.1101/2020.03.16.992933 ◽

2020 ◽

Cited By ~ 1

Author(s):

Valentine Murigneux ◽

Subash Kumar Rai ◽

Agnelo Furtado ◽

Timothy J.C. Bruxner ◽

Wei Tian ◽

...

Keyword(s):

De Novo ◽

Cost Effective ◽

Genome Project ◽

Plant Genome ◽

Sequencing Data ◽

Sequencing Technologies ◽

Oxford Nanopore ◽

Long Read ◽

The Cost ◽

Genome Assemblies

AbstractSequencing technologies have advanced to the point where it is possible to generate high accuracy, haplotype resolved, chromosome scale assemblies. Several long read sequencing technologies are available on the market and a growing number of algorithms have been developed over the last years to assemble the reads generated by those technologies. When starting a new genome project, it is therefore challenging to select the most cost-effective sequencing technology as well as the most appropriate software for assembly and polishing. For this reason, it is important to benchmark different approaches applied to the same sample. Here, we report a comparison of three long read sequencing technologies applied to the de novo assembly of a plant genome, Macadamia jansenii. We have generated sequencing data using Pacific Biosciences (Sequel I), Oxford Nanopore Technologies (PromethION) and BGI (single-tube Long Fragment Read) technologies for the same sample. Several assemblers were benchmarked in the assembly of PacBio and Nanopore reads. Results obtained from combining long read technologies or short read and long read technologies are also presented. The assemblies were compared for contiguity, accuracy and completeness as well as sequencing costs and DNA material requirements. Overall, the three long read technologies produced highly contiguous and complete genome assemblies of Macadamia jansenii. At the time of sequencing, the cost associated with each method was significantly different but continuous improvements in technologies have resulted in greater accuracy, increased throughput and reduced costs. We propose updating this comparison regularly with reports on significant iterations of the sequencing technologies.

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Comprehensive identification of transposable element insertions using multiple sequencing technologies

Nature Communications ◽

10.1038/s41467-021-24041-8 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Chong Chu ◽

Rebeca Borges-Monroy ◽

Vinayak V. Viswanadham ◽

Soohyun Lee ◽

Heng Li ◽

...

Keyword(s):

Transposable Element ◽

Structure And Function ◽

Endogenous Retroviruses ◽

Whole Genome Sequencing Data ◽

Whole Genome ◽

Sequencing Data ◽

Short Read ◽

Sequencing Technologies ◽

Long Read ◽

And Function

AbstractTransposable elements (TEs) help shape the structure and function of the human genome. When inserted into some locations, TEs may disrupt gene regulation and cause diseases. Here, we present xTea (x-Transposable element analyzer), a tool for identifying TE insertions in whole-genome sequencing data. Whereas existing methods are mostly designed for short-read data, xTea can be applied to both short-read and long-read data. Our analysis shows that xTea outperforms other short read-based methods for both germline and somatic TE insertion discovery. With long-read data, we created a catalogue of polymorphic insertions with full assembly and annotation of insertional sequences for various types of retroelements, including pseudogenes and endogenous retroviruses. Notably, we find that individual genomes have an average of nine groups of full-length L1s in centromeres, suggesting that centromeres and other highly repetitive regions such as telomeres are a significant yet unexplored source of active L1s. xTea is available at https://github.com/parklab/xTea.

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

Microbiology Resource Announcements ◽

10.1128/mra.00818-19 ◽

2019 ◽

Vol 8 (34) ◽

Cited By ~ 1

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.

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Identifying contamination with advanced visualization and analysis practices: metagenomic approaches for eukaryotic genome assemblies

PeerJ ◽

10.7717/peerj.1839 ◽

2016 ◽

Vol 4 ◽

pp. e1839 ◽

Cited By ~ 57

Author(s):

Tom O. Delmont ◽

A. Murat Eren

Keyword(s):

High Throughput Sequencing ◽

Draft Genome ◽

Cost Effective ◽

Single Copy ◽

Eukaryotic Genome ◽

Sequencing Data ◽

Bacterial Genomes ◽

Long Read ◽

Domains Of Life ◽

Genome Assemblies

High-throughput sequencing provides a fast and cost-effective mean to recover genomes of organisms from all domains of life. However, adequate curation of the assembly results against potential contamination of non-target organisms requires advanced bioinformatics approaches and practices. Here, we re-analyzed the sequencing data generated for the tardigradeHypsibius dujardini,and created a holistic display of the eukaryotic genome assembly using DNA data originating from two groups and eleven sequencing libraries. By using bacterial single-copy genes, k-mer frequencies, and coverage values of scaffolds we could identify and characterize multiple near-complete bacterial genomes from the raw assembly, and curate a 182 Mbp draft genome forH. dujardinisupported by RNA-Seq data. Our results indicate that most contaminant scaffolds were assembled from Moleculo long-read libraries, and most of these contaminants have differed between library preparations. Our re-analysis shows that visualization and curation of eukaryotic genome assemblies can benefit from tools designed to address the needs of today’s microbiologists, who are constantly challenged by the difficulties associated with the identification of distinct microbial genomes in complex environmental metagenomes.

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SVJedi: Genotyping structural variations with long reads

10.1101/849208 ◽

2019 ◽

Author(s):

Lolita Lecompte ◽

Pierre Peterlongo ◽

Dominique Lavenier ◽

Claire Lemaitre

Keyword(s):

Sequencing Data ◽

Structural Variations ◽

Short Read ◽

Sequencing Technologies ◽

Long Reads ◽

Oxford Nanopore ◽

Clinical Diagnoses ◽

Long Read ◽

Reference Sequences ◽

The One

AbstractMotivationStudies on structural variants (SV) are expanding rapidly. As a result, and thanks to third generation sequencing technologies, the number of discovered SVs is increasing, especially in the human genome. At the same time, for several applications such as clinical diagnoses, it is important to genotype newly sequenced individuals on well defined and characterized SVs. Whereas several SV genotypers have been developed for short read data, there is a lack of such dedicated tool to assess whether known SVs are present or not in a new long read sequenced sample, such as the one produced by Pacific Biosciences or Oxford Nanopore Technologies.ResultsWe present a novel method to genotype known SVs from long read sequencing data. The method is based on the generation of a set of reference sequences that represent the two alleles of each structural variant. Long reads are aligned to these reference sequences. Alignments are then analyzed and filtered out to keep only informative ones, to quantify and estimate the presence of each SV allele and the allele frequencies. We provide an implementation of the method, SVJedi, to genotype insertions and deletions with long reads. The tool has been applied to both simulated and real human datasets and achieves high genotyping accuracy. We also demonstrate that SV genotyping is considerably improved with SVJedi compared to other approaches, namely SV discovery and short read SV genotyping approaches.Availabilityhttps://github.com/llecompte/[email protected]

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LongStitch: High-quality genome assembly correction and scaffolding using long reads

10.1101/2021.06.17.448848 ◽

2021 ◽

Author(s):

Lauren Coombe ◽

Janet X Li ◽

Theodora Lo ◽

Johnathan Wong ◽

Vladimir Nikolic ◽

...

Keyword(s):

Genome Assembly ◽

De Novo ◽

Draft Genome ◽

Model Organisms ◽

High Quality ◽

De Novo Genome Assembly ◽

Long Reads ◽

Long Read ◽

Genomic Regions ◽

Genome Assemblies

Background Generating high-quality de novo genome assemblies is foundational to the genomics study of model and non-model organisms. In recent years, long-read sequencing has greatly benefited genome assembly and scaffolding, a process by which assembled sequences are ordered and oriented through the use of long-range information. Long reads are better able to span repetitive genomic regions compared to short reads, and thus have tremendous utility for resolving problematic regions and helping generate more complete draft assemblies. Here, we present LongStitch, a scalable pipeline that corrects and scaffolds draft genome assemblies exclusively using long reads. Results LongStitch incorporates multiple tools developed by our group and runs in up to three stages, which includes initial assembly correction (Tigmint-long), followed by two incremental scaffolding stages (ntLink and ARKS-long). Tigmint-long and ARKS-long are misassembly correction and scaffolding utilities, respectively, previously developed for linked reads, that we adapted for long reads. Here, we describe the LongStitch pipeline and introduce our new long-read scaffolder, ntLink, which utilizes lightweight minimizer mappings to join contigs. LongStitch was tested on short and long-read assemblies of three different human individuals using corresponding nanopore long-read data, and improves the contiguity of each assembly from 2.0-fold up to 304.6-fold (as measured by NGA50 length). Furthermore, LongStitch generates more contiguous and correct assemblies compared to state-of-the-art long-read scaffolder LRScaf in most tests, and consistently runs in under five hours using less than 23GB of RAM. Conclusions Due to its effectiveness and efficiency in improving draft assemblies using long reads, we expect LongStitch to benefit a wide variety of de novo genome assembly projects. The LongStitch pipeline is freely available at https://github.com/bcgsc/longstitch.

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Clustering de Novo by Gene of Long Reads from Transcriptomics Data

10.1101/170035 ◽

2017 ◽

Cited By ~ 3

Author(s):

Camille Marchet ◽

Lolita Lecompte ◽

Corinne Da Silva ◽

Corinne Cruaud ◽

Jean-Marc Aury ◽

...

Keyword(s):

De Novo ◽

Free Access ◽

Sequencing Data ◽

Base Pairs ◽

Long Reads ◽

Oxford Nanopore ◽

Processing Step ◽

Whole Transcriptome Sequencing ◽

Long Read ◽

Transcriptomics Data

AbstractLong-read sequencing currently provides sequences of several thousand base pairs. This allows to obtain complete transcripts, which offers an un-precedented vision of the cellular transcriptome.However the literature is lacking tools to cluster such data de novo, in particular for Oxford Nanopore Technologies reads, because of the inherent high error rate compared to short reads.Our goal is to process reads from whole transcriptome sequencing data accurately and without a reference genome in order to reliably group reads coming from the same gene. This de novo approach is therefore particularly suitable for non-model species, but can also serve as a useful pre-processing step to improve read mapping. Our contribution is both to propose a new algorithm adapted to clustering of reads by gene and a practical and free access tool that permits to scale the complete processing of eukaryotic transcriptomes.We sequenced a mouse RNA sample using the MinION device, this dataset is used to compare our solution to other algorithms used in the context of biological clustering. We demonstrate its is better-suited for transcriptomics long reads. When a reference is available thus mapping possible, we show that it stands as an alternative method that predicts complementary clusters.

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Extensive genomic and transcriptomic variation defines the chromosome-scale assembly of Haemonchus contortus, a model gastrointestinal worm

10.1101/2020.02.18.945246 ◽

2020 ◽

Cited By ~ 2

Author(s):

Stephen R. Doyle ◽

Alan Tracey ◽

Roz Laing ◽

Nancy Holroyd ◽

David Bartley ◽

...

Keyword(s):

Genome Assembly ◽

Haemonchus Contortus ◽

Vaccine Development ◽

De Novo ◽

Anthelmintic Resistance ◽

Draft Genome ◽

Small Ruminants ◽

High Quality ◽

Long Read ◽

Genome Assemblies

AbstractBackgroundHaemonchus contortus is a globally distributed and economically important gastrointestinal pathogen of small ruminants, and has become the key nematode model for studying anthelmintic resistance and other parasite-specific traits among a wider group of parasites including major human pathogens. Two draft genome assemblies for H. contortus were reported in 2013, however, both were highly fragmented, incomplete, and differed from one another in important respects. While the introduction of long-read sequencing has significantly increased the rate of production and contiguity of de novo genome assemblies broadly, achieving high quality genome assemblies for small, genetically diverse, outcrossing eukaryotic organisms such as H. contortus remains a significant challenge.ResultsHere, we report using PacBio long read and OpGen and 10X Genomics long-molecule methods to generate a highly contiguous 283.4 Mbp chromosome-scale genome assembly including a resolved sex chromosome. We show a remarkable pattern of almost complete conservation of chromosome content (synteny) with Caenorhabditis elegans, but almost no conservation of gene order. Long-read transcriptome sequence data has allowed us to define coordinated transcriptional regulation throughout the life cycle of the parasite, and refine our understanding of cis- and trans-splicing relative to that observed in C. elegans. Finally, we use this assembly to give a comprehensive picture of chromosome-wide genetic diversity both within a single isolate and globally.ConclusionsThe H. contortus MHco3(ISE).N1 genome assembly presented here represents the most contiguous and resolved nematode assembly outside of the Caenorhabditis genus to date, together with one of the highest-quality set of predicted gene features. These data provide a high-quality comparison for understanding the evolution and genomics of Caenorhabditis and other nematodes, and extends the experimental tractability of this model parasitic nematode in understanding pathogen biology, drug discovery and vaccine development, and important adaptive traits such as drug resistance.

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Complete Genome Sequence of Vibrio rotiferianus Strain AM7

Microbiology Resource Announcements ◽

10.1128/mra.01591-19 ◽

2020 ◽

Vol 9 (21) ◽

Author(s):

Kentaro Miyazaki ◽

Apirak Wiseschart ◽

Kusol Pootanakit ◽

Kei Kitahara

Keyword(s):

Genome Sequence ◽

Complete Genome Sequence ◽

Complete Genome ◽

The Novel ◽

Sequencing Data ◽

Short Read ◽

Content Type ◽

Short Read Sequencing ◽

Oxford Nanopore ◽

Long Read

ABSTRACT We isolated the novel strain Vibrio rotiferianus AM7 from the shell of an abalone. In this article, we report the complete genome sequence of this organism, which was obtained by combining Oxford Nanopore long-read and Illumina short-read sequencing data.

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