scholarly journals Metagenomic data for Halichondria panicea from Illumina and Nanopore sequencing and preliminary genome assemblies for the sponge and two microbial symbionts.

2021 ◽  
Author(s):  
Brian W Strehlow ◽  
Astrid Schuster ◽  
Warren R Francis ◽  
Donald E Canfield
Keyword(s):  
Additional Data ◽  
Illumina Miseq ◽  
Metagenomic Data ◽  
Single Individual ◽  
Halichondria Panicea ◽  
Oxford Nanopore ◽  
Long Read ◽  
Microbial Symbionts ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

Objectives: These data were collected to generate a novel reference metagenome for the sponge Halichondria panicea and its microbiome for subsequent differential expression analyses. Data description: These data include raw sequences from four separate sequencing runs of the metagenome of a single individual of H. panicea - one Illumina MiSeq (2x300 bp, paired-end) run and three Oxford Nanopore Technologies (ONT) long-read sequencing runs, generating 53.8 and 7.42 Gbp respectively. Comparing assemblies of Illumina, ONT and an Illumina-ONT hybrid revealed the hybrid to be the best assembly, comprising 163 Mbp in 63,555 scaffolds (N50: 3,084). This assembly, however, was still highly fragmented and only contained 52% of core metazoan genes (with 77.9% partial genes), so it was also not complete. However, this sponge is an emerging model species for field and laboratory work, and there is considerable interest in genomic sequencing of this species. Although the resultant assemblies from the data presented here are suboptimal, this data note can inform future studies by providing an estimated genome size and coverage requirements for future sequencing, sharing additional data to potentially improve other suboptimal assemblies of this species, and outlining potential limitations and pitfalls of the combined Illumina and ONT approach to novel genome sequencing.

scholarly journals Chasing perfection: validation and polishing strategies for telomere-to-telomere genome assemblies

2021 ◽  
Author(s):  
Arang Rhie ◽  
Ann Mc Cartney ◽  
Kishwar Shafin ◽  
Michael Alonge ◽  
Andrey Bzikadze ◽  
...  
Keyword(s):  
Genome Assembly ◽  
Tandem Repeats ◽  
Hydatidiform Mole ◽  
Segmental Duplications ◽  
Sequencing Technologies ◽  
Oxford Nanopore ◽  
Human Genome Assembly ◽  
Long Read ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

Abstract Advances in long-read sequencing technologies and genome assembly methods have enabled the recent completion of the first Telomere-to-Telomere (T2T) human genome assembly, which resolves complex segmental duplications and large tandem repeats, including centromeric satellite arrays in a complete hydatidiform mole (CHM13). Though derived from highly accurate sequencing, evaluation revealed that the initial T2T draft assembly had evidence of small errors and structural misassemblies. To correct these errors, we designed a novel repeat-aware polishing strategy that made accurate assembly corrections in large repeats without overcorrection, ultimately fixing 51% of the existing errors and improving the assembly QV to 73.9. By comparing our results to standard automated polishing tools, we outline common polishing errors and offer practical suggestions for genome projects with limited resources. We also show how sequencing biases in both PacBio HiFi and Oxford Nanopore Technologies reads cause signature assembly errors that can be corrected with a diverse panel of sequencing technologies

scholarly journals Comparison of the two up-to-date sequencing technologies for genome assembly: HiFi reads of Pacbio Sequel II system and ultralong reads of Oxford Nanopore

2020 ◽  
Author(s):  
Dandan Lang ◽  
Shilai Zhang ◽  
Pingping Ren ◽  
Fan Liang ◽  
Zongyi Sun ◽  
...  
Keyword(s):  
Gene Families ◽  
Single Chromosome ◽  
Small Indels ◽  
Base Level ◽  
Sequencing Technologies ◽  
Oxford Nanopore ◽  
Long Read ◽  
Single Rice ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

AbstractThe availability of reference genomes has revolutionized the study of biology. Multiple competing technologies have been developed to improve the quality and robustness of genome assemblies during the last decade. The two widely-used long read sequencing providers – Pacbio (PB) and Oxford Nanopore Technologies (ONT) – have recently updated their platforms: PB enable high throughput HiFi reads with base-level resolution with >99% and ONT generated reads as long as 2 Mb. We applied the two up-to-date platforms to one single rice individual, and then compared the two assemblies to investigate the advantages and limitations of each. The results showed that ONT ultralong reads delivered higher contiguity producing a total of 18 contigs of which 10 were assembled into a single chromosome compared to that of 394 contigs and three chromosome-level contigs for the PB assembly. The ONT ultralong reads also prevented assembly errors caused by long repetitive regions for which we observed a total 44 genes of false redundancies and 10 genes of false losses in the PB assembly leading to over/under-estimations of the gene families in those long repetitive regions. We also noted that the PB HiFi reads generated assemblies with considerably less errors at the level of single nucleotide and small InDels than that of the ONT assembly which generated an average 1.06 errors per Kb assembly and finally engendered 1,475 incorrect gene annotations via altered or truncated protein predictions.

scholarly journals De-novo Assembly of Limnospira fusiformis Using Ultra-Long Reads

2021 ◽  
Vol 12 ◽  
Author(s):  
McKenna Hicks ◽  
Thuy-Khanh Tran-Dao ◽  
Logan Mulroney ◽  
David L. Bernick
Keyword(s):  
Phylogenetic Analysis ◽  
Type Strain ◽  
Reference Genome ◽  
De Novo ◽  
Illumina Miseq ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Oxford Nanopore Technologies ◽  
Rdna Analysis

The Limnospira genus is a recently established clade that is economically important due to its worldwide use in biotechnology and agriculture. This genus includes organisms that were reclassified from Arthrospira, which are commercially marketed as “Spirulina.” Limnospira are photoautotrophic organisms that are widely used for research in nutrition, medicine, bioremediation, and biomanufacturing. Despite its widespread use, there is no closed genome for the Limnospira genus, and no reference genome for the type strain, Limnospira fusiformis. In this work, the L. fusiformis genome was sequenced using Oxford Nanopore Technologies MinION and assembled using only ultra-long reads (>35 kb). This assembly was polished with Illumina MiSeq reads sourced from an axenic L. fusiformis culture; axenicity was verified via microscopy and rDNA analysis. Ultra-long read sequencing resulted in a 6.42 Mb closed genome assembled as a single contig with no plasmid. Phylogenetic analysis placed L. fusiformis in the Limnospira clade; some Arthrospira were also placed in this clade, suggesting a misclassification of these strains. This work provides a fully closed and accurate reference genome for the economically important type strain, L. fusiformis. We also present a rapid axenicity method to isolate L. fusiformis. These contributions enable future biotechnological development of L. fusiformis by way of genetic engineering.

scholarly journals Complete and Circularized Genome Assemblies of the Kroppenstedtia eburnea Genus Type Strain and the Kroppenstedtia pulmonis Species Type Strain with MiSeq and MinION Sequence Data

2020 ◽  
Vol 9 (44) ◽  
Author(s):  
Christopher A. Gulvik ◽  
Dhwani Batra ◽  
Lori A. Rowe ◽  
Milli Sheth ◽  
Ben W. Humrighouse ◽  
...  
Keyword(s):  
Type Strain ◽  
Sequence Data ◽  
Gene Clusters ◽  
Illumina Miseq ◽  
Putative Gene ◽  
Content Type ◽  
Oxford Nanopore ◽  
Species Type ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

ABSTRACT Kroppenstedtia eburnea DSM 45196T and Kroppenstedtia pulmonis W9323T are aerobic, Gram-positive, filamentous, chemoorganotrophic thermoactinomycetes. Here, we report on the complete and circular genome assemblies generated using Illumina MiSeq and Oxford Nanopore Technologies MinION reads. Putative gene clusters predicted to be involved in the production of secondary metabolites were also identified.

scholarly journals Sequencing and Chromosome-Scale Assembly of Plant Genomes, Brassica rapa as a Use Case

Biology ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 732
Author(s):  
Benjamin Istace ◽  
Caroline Belser ◽  
Cyril Falentin ◽  
Karine Labadie ◽  
Franz Boideau ◽  
...  
Keyword(s):  
Long Range ◽  
Brassica Rapa ◽  
Plant Genome ◽  
Use Case ◽  
Computer Algorithms ◽  
High Quality ◽  
Oxford Nanopore ◽  
Long Read ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

With the rise of long-read sequencers and long-range technologies, delivering high-quality plant genome assemblies is no longer reserved to large consortia. Not only sequencing techniques, but also computer algorithms have reached a point where the reconstruction of assemblies at the chromosome scale is now feasible at the laboratory scale. Current technologies, in particular long-range technologies, are numerous, and selecting the most promising one for the genome of interest is crucial to obtain optimal results. In this study, we resequenced the genome of the yellow sarson, Brassica rapa cv. Z1, using the Oxford Nanopore PromethION sequencer and assembled the sequenced data using current assemblers. To reconstruct complete chromosomes, we used and compared three long-range scaffolding techniques, optical mapping, Omni-C, and Pore-C sequencing libraries, commercialized by Bionano Genomics, Dovetail Genomics, and Oxford Nanopore Technologies, respectively, or a combination of the three, in order to evaluate the capability of each technology.

scholarly journals Sequencing and Chromosome-Scale Assembly of Plant Genomes, Brassica rapa as a Use Case

2021 ◽  
Author(s):  
Benjamin Istace ◽  
Caroline Belser ◽  
Cyril Falentin ◽  
Karine Labadie ◽  
Franz Boideau ◽  
...  
Keyword(s):  
Long Range ◽  
Brassica Rapa ◽  
Plant Genome ◽  
Use Case ◽  
Computer Algorithms ◽  
High Quality ◽  
Oxford Nanopore ◽  
Long Read ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

With the rise of long-read sequencers and long-range technologies, delivering high-quality plant genome assemblies is no longer reserved to large consortia. Indeed, sequencing techniques but also computer algorithms have reached a point where the reconstruction of assemblies at the chromosome-scale is now feasible at the laboratory scale. Current technologies, and especially long-range technologies, are numerous and selecting the most promising one for the genome of interest is crucial to obtain optimal results. In this study, we resequenced the genome of the yellow sarson, Brassica rapa cv. Z1, using the Oxford Nanopore PromethION sequencer and assembled the sequenced data using current assemblers. To reconstruct complete chromosomes, we used and compared three long-range techniques, optical mapping, Omni-C and Pore-C sequencing libraries commercialized by Bionano Genomics, Dovetail Genomics and Oxford Nanopore Technologies respectively, or a combination of the three, in order to evaluate the capability of each technology.

scholarly journals Chasing perfection: validation and polishing strategies for telomere-to-telomere genome assemblies

2021 ◽  
Author(s):  
Ann M Mc Cartney ◽  
Kishwar Shafin ◽  
Michael Alonge ◽  
Andrey V Bzikadze ◽  
Giulio Formenti ◽  
...  
Keyword(s):  
Genome Assembly ◽  
Tandem Repeats ◽  
Hydatidiform Mole ◽  
Segmental Duplications ◽  
Sequencing Technologies ◽  
Oxford Nanopore ◽  
Human Genome Assembly ◽  
Long Read ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

Advances in long-read sequencing technologies and genome assembly methods have enabled the recent completion of the first Telomere-to-Telomere (T2T) human genome assembly, which resolves complex segmental duplications and large tandem repeats, including centromeric satellite arrays in a complete hydatidiform mole (CHM13). Though derived from highly accurate sequencing, evaluation revealed that the initial T2T draft assembly had evidence of small errors and structural misassemblies. To correct these errors, we designed a novel repeat-aware polishing strategy that made accurate assembly corrections in large repeats without overcorrection, ultimately fixing 51% of the existing errors and improving the assembly QV to 73.9. By comparing our results to standard automated polishing tools, we outline common polishing errors and offer practical suggestions for genome projects with limited resources. We also show how sequencing biases in both PacBio HiFi and Oxford Nanopore Technologies reads cause signature assembly errors that can be corrected with a diverse panel of sequencing technologies.

scholarly journals Hapo-G, haplotype-aware polishing of genome assemblies with accurate reads

2021 ◽  
Vol 3 (2) ◽  
Author(s):  
Jean-Marc Aury ◽  
Benjamin Istace
Keyword(s):  
Single Molecule ◽  
Direct Consequence ◽  
High Quality ◽  
Sequencing Errors ◽  
Coding Regions ◽  
Sequencing Technologies ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Genome Assemblies

Abstract Single-molecule sequencing technologies have recently been commercialized by Pacific Biosciences and Oxford Nanopore with the promise of sequencing long DNA fragments (kilobases to megabases order) and then, using efficient algorithms, provide high quality assemblies in terms of contiguity and completeness of repetitive regions. However, the error rate of long-read technologies is higher than that of short-read technologies. This has a direct consequence on the base quality of genome assemblies, particularly in coding regions where sequencing errors can disrupt the coding frame of genes. In the case of diploid genomes, the consensus of a given gene can be a mixture between the two haplotypes and can lead to premature stop codons. Several methods have been developed to polish genome assemblies using short reads and generally, they inspect the nucleotide one by one, and provide a correction for each nucleotide of the input assembly. As a result, these algorithms are not able to properly process diploid genomes and they typically switch from one haplotype to another. Herein we proposed Hapo-G (Haplotype-Aware Polishing Of Genomes), a new algorithm capable of incorporating phasing information from high-quality reads (short or long-reads) to polish genome assemblies and in particular assemblies of diploid and heterozygous genomes.

Dual Isoform Sequencing Reveals a Multifaceted Transcriptional Architecture of a Prototype Baculovirus

2021 ◽  
Author(s):  
Gábor Torma ◽  
Dóra Tombácz ◽  
Norbert Moldován ◽  
Ádám Fülöp ◽  
István Prazsák ◽  
...  
Keyword(s):  
Protein Coding ◽  
Rna Molecules ◽  
Non Coding Rna ◽  
Oxford Nanopore ◽  
The Pacific ◽  
Viral Genes ◽  
Long Read ◽  
Oxford Nanopore Technologies ◽  
Overlapping Transcripts

Abstract In this study, we used two long-read sequencing (LRS) techniques, Sequel from the Pacific Biosciences and MinION from Oxford Nanopore Technologies, for the transcriptional characterization of a prototype baculovirus, Autographacalifornica multiple nucleopolyhedrovirus. LRS is able to read full-length RNA molecules, and thereby to distinguish between transcript isoforms, mono- and polycistronic RNAs, and overlapping transcripts. Altogether, we detected 875 transcripts, of which 759 are novel and 116 have been annotated previously. These RNA molecules include 41 novel putative protein coding transcript (each containing 5’-truncated in-frame ORFs), 14 monocistronic transcripts, 99 multicistronic RNAs, 101 non-coding RNA, and 504 length isoforms. We also detected RNA methylation in 12 viral genes and RNA hyper-editing in the longer 5’-UTR transcript isoform of ORF 19 gene.

scholarly journals Microbial diversity characterization of seawater in a pilot study using Oxford Nanopore Technologies long-read sequencing

2020 ◽  
Author(s):  
Michael Liem ◽  
Tonny Regensburg-Tuïnk ◽  
Christiaan Henkel ◽  
Hans Jansen ◽  
Herman Spaink
Keyword(s):  
Microbial Diversity ◽  
Environmental Samples ◽  
Sea Water ◽  
Flow Cells ◽  
Oxford Nanopore ◽  
Challenging Tasks ◽  
Long Read ◽  
Close Relatives ◽  
Oxford Nanopore Technologies

Abstract Objective: Currently the majority of non-culturable microbes in sea water are yet to be discovered, Nanopore offers a solution to overcome the challenging tasks to identify the genomes and complex composition of oceanic microbiomes. In this study we evaluate the utility of Oxford Nanopore Technologies (ONT) sequencing to characterize microbial diversity in seawater from multiple locations. We compared the microbial species diversity of retrieved environmental samples from two different locations and time points.Results: With only three ONT flow cells we were able to identify thousands of organisms, including bacteriophages, from which a large part at species level. It was possible to assemble genomes from environmental samples with Flye. In several cases this resulted in >1 Mbp contigs and in the particular case of a Thioglobus singularis species it even produced a near complete genome. k-mer analysis reveals that a large part of the data represents species of which close relatives have not yet been deposited to the database. These results show that our approach is suitable for scalable genomic investigations such as monitoring oceanic biodiversity and provides a new platform for education in biodiversity.

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