scholarly journals Platanus_B: an accurate de novo assembler for bacterial genomes using an iterative error-removal process

DNA Research ◽  
2020 ◽  
Vol 27 (3) ◽  
Author(s):  
Rei Kajitani ◽  
Dai Yoshimura ◽  
Yoshitoshi Ogura ◽  
Yasuhiro Gotoh ◽  
Tetsuya Hayashi ◽  
...  
Keyword(s):  
High Resolution ◽  
Large Scale ◽  
De Novo ◽  
Bacterial Genomes ◽  
Long Reads ◽  
Wide Range ◽  
Long Read ◽  
Phylogenomic Analyses ◽  
Error Removal ◽  
Iterative Error

Abstract De novo assembly of short DNA reads remains an essential technology, especially for large-scale projects and high-resolution variant analyses in epidemiology. However, the existing tools often lack sufficient accuracy required to compare closely related strains. To facilitate such studies on bacterial genomes, we developed Platanus_B, a de novo assembler that employs iterations of multiple error-removal algorithms. The benchmarks demonstrated the superior accuracy and high contiguity of Platanus_B, in addition to its ability to enhance the hybrid assembly of both short and nanopore long reads. Although the hybrid strategies for short and long reads were effective in achieving near full-length genomes, we found that short-read-only assemblies generated with Platanus_B were sufficient to obtain ≥90% of exact coding sequences in most cases. In addition, while nanopore long-read-only assemblies lacked fine-scale accuracies, inclusion of short reads was effective in improving the accuracies. Platanus_B can, therefore, be used for comprehensive genomic surveillances of bacterial pathogens and high-resolution phylogenomic analyses of a wide range of bacteria.

scholarly journals Deep repeat resolution—the assembly of the Drosophila Histone Complex

2018 ◽  
Vol 47 (3) ◽  
pp. e18-e18 ◽  
Author(s):  
Philipp Bongartz ◽  
Siegfried Schloissnig
Keyword(s):  
De Novo ◽  
Machine Learning Algorithms ◽  
Single Nucleotide Variants ◽  
Major Step ◽  
Base Pairs ◽  
Sequencing Technologies ◽  
Long Reads ◽  
Wide Range ◽  
Long Read ◽  
Genome Assemblies

Abstract Though the advent of long-read sequencing technologies has led to a leap in contiguity of de novo genome assemblies, current reference genomes of higher organisms still do not provide unbroken sequences of complete chromosomes. Despite reads in excess of 30 000 base pairs, there are still repetitive structures that cannot be resolved by current state-of-the-art assemblers. The most challenging of these structures are tandemly arrayed repeats, which occur in the genomes of all eukaryotes. Untangling tandem repeat clusters is exceptionally difficult, since the rare differences between repeat copies are obscured by the high error rate of long reads. Solving this problem would constitute a major step towards computing fully assembled genomes. Here, we demonstrate by example of the Drosophila Histone Complex that via machine learning algorithms, it is possible to exploit the underlying distinguishing patterns of single nucleotide variants of repeats from very noisy data to resolve a large and highly conserved repeat cluster. The ideas explored in this paper are a first step towards the automated assembly of complex repeat structures and promise to be applicable to a wide range of eukaryotic genomes.

scholarly journals Accurate long-read de novo assembly evaluation with Inspector

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.

scholarly journals Dependence of Model Energy Spectra on Vertical Resolution

2016 ◽  
Vol 144 (4) ◽  
pp. 1407-1421 ◽  
Author(s):  
Michael L. Waite
Keyword(s):  
High Resolution ◽  
Large Scale ◽  
Vertical Mixing ◽  
Energy Spectra ◽  
Small Scale ◽  
Rossby Number ◽  
Vertical Resolution ◽  
Wide Range ◽  
Qualitative Shape ◽  
Vertical Grid

Abstract Many high-resolution atmospheric models can reproduce the qualitative shape of the atmospheric kinetic energy spectrum, which has a power-law slope of −3 at large horizontal scales that shallows to approximately −5/3 in the mesoscale. This paper investigates the possible dependence of model energy spectra on the vertical grid resolution. Idealized simulations forced by relaxation to a baroclinically unstable jet are performed for a wide range of vertical grid spacings Δz. Energy spectra are converged for Δz 200 m but are very sensitive to resolution with 500 m ≤ Δz ≤ 2 km. The nature of this sensitivity depends on the vertical mixing scheme. With no vertical mixing or with weak, stability-dependent mixing, the mesoscale spectra are artificially amplified by low resolution: they are shallower and extend to larger scales than in the converged simulations. By contrast, vertical hyperviscosity with fixed grid-scale damping rate has the opposite effect: underresolved spectra are spuriously steepened. High-resolution spectra are converged except for the stability-dependent mixing case, which are damped by excessive mixing due to enhanced shear over a wide range of horizontal scales. It is shown that converged spectra require resolution of all vertical scales associated with the resolved horizontal structures: these include quasigeostrophic scales for large-scale motions with small Rossby number and the buoyancy scale for small-scale motions at large Rossby number. It is speculated that some model energy spectra may be contaminated by low vertical resolution, and it is recommended that vertical-resolution sensitivity tests always be performed.

scholarly journals A Universal, Genomewide GuideFinder for CRISPR/Cas9 Targeting in Microbial Genomes

mSphere ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Michelle Spoto ◽  
Changhui Guan ◽  
Elizabeth Fleming ◽  
Julia Oh
Keyword(s):  
Gene Function ◽  
Large Scale ◽  
Essential Gene ◽  
Bacterial Species ◽  
Bacterial Genome ◽  
Model Organisms ◽  
Design Parameters ◽  
Bacterial Genomes ◽  
Wide Range ◽  
User Friendly

ABSTRACT The CRISPR/Cas system has significant potential to facilitate gene editing in a variety of bacterial species. CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa) represent modifications of the CRISPR/Cas9 system utilizing a catalytically inactive Cas9 protein for transcription repression and activation, respectively. While CRISPRi and CRISPRa have tremendous potential to systematically investigate gene function in bacteria, few programs are specifically tailored to identify guides in draft bacterial genomes genomewide. Furthermore, few programs offer open-source code with flexible design parameters for bacterial targeting. To address these limitations, we created GuideFinder, a customizable, user-friendly program that can design guides for any annotated bacterial genome. GuideFinder designs guides from NGG protospacer-adjacent motif (PAM) sites for any number of genes by the use of an annotated genome and FASTA file input by the user. Guides are filtered according to user-defined design parameters and removed if they contain any off-target matches. Iteration with lowered parameter thresholds allows the program to design guides for genes that did not produce guides with the more stringent parameters, one of several features unique to GuideFinder. GuideFinder can also identify paired guides for targeting multiplicity, whose validity we tested experimentally. GuideFinder has been tested on a variety of diverse bacterial genomes, finding guides for 95% of genes on average. Moreover, guides designed by the program are functionally useful—focusing on CRISPRi as a potential application—as demonstrated by essential gene knockdown in two staphylococcal species. Through the large-scale generation of guides, this open-access software will improve accessibility to CRISPR/Cas studies of a variety of bacterial species. IMPORTANCE With the explosion in our understanding of human and environmental microbial diversity, corresponding efforts to understand gene function in these organisms are strongly needed. CRISPR/Cas9 technology has revolutionized interrogation of gene function in a wide variety of model organisms. Efficient CRISPR guide design is required for systematic gene targeting. However, existing tools are not adapted for the broad needs of microbial targeting, which include extraordinary species and subspecies genetic diversity, the overwhelming majority of which is characterized by draft genomes. In addition, flexibility in guide design parameters is important to consider the wide range of factors that can affect guide efficacy, many of which can be species and strain specific. We designed GuideFinder, a customizable, user-friendly program that addresses the limitations of existing software and that can design guides for any annotated bacterial genome with numerous features that facilitate guide design in a wide variety of microorganisms.

scholarly journals Impact of Chimera-less Long Reads on Metagenomics of Human Gut Viromes Treated With Multiple Displacement Amplification

2020 ◽  
Author(s):  
Yuya Kiguchi ◽  
Suguru Nishijima ◽  
Naveen Kumar ◽  
Masahira Hattori ◽  
Wataru Suda
Keyword(s):  
De Novo ◽  
Multiple Displacement Amplification ◽  
Read Length ◽  
Human Gut ◽  
Average Read Length ◽  
Sequencing Technologies ◽  
Genomic Complexity ◽  
Long Reads ◽  
Long Read ◽  
The Impact

Abstract Background: The ecological and biological features of the indigenous phage community (virome) in the human gut microbiome are poorly understood, possibly due to many fragmented contigs and fewer complete genomes based on conventional short-read metagenomics. Long-read sequencing technologies have attracted attention as an alternative approach to reconstruct long and accurate contigs from microbial communities. However, the impact of long-read metagenomics on human gut virome analysis has not been well evaluated. Results: Here we present chimera-less PacBio long-read metagenomics of multiple displacement amplification (MDA)-treated human gut virome DNA. The method included the development of a novel bioinformatics tool, SACRA (Split Amplified Chimeric Read Algorithm), which efficiently detects and splits numerous chimeric reads in PacBio reads from the MDA-treated virome samples. SACRA treatment of PacBio reads from five samples markedly reduced the average chimera ratio from 72 to 1.5%, generating chimera-less PacBio reads with an average read-length of 1.8 kb. De novo assembly of the chimera-less long reads generated contigs with an average N50 length of 11.1 kb, whereas those of MiSeq short reads from the same samples were 0.7 kb, dramatically improving contig extension. Alignment of both contig sets generated 378 high-quality merged contigs (MCs) composed of the minimum scaffolds of 434 MiSeq and 637 PacBio contigs, respectively, and also identified numerous MiSeq short fragmented contigs ≤500 bp additionally aligned to MCs, which possibly originated from a small fraction of MiSeq chimeric reads. The alignment also revealed that fragmentations of the scaffolded MiSeq contigs were caused primarily by genomic complexity of the community, including local repeats, hypervariable regions, and highly conserved sequences in and between the phage genomes. We identified 142 complete and near-complete phage genomes including 108 novel genomes, varying from 5 to 185 kb in length, the majority of which were predicted to be Microviridae phages including several variants with homologous but distinct genomes, which were fragmented in MiSeq contigs. Conclusions: Long-read metagenomics coupled with SACRA provides an improved method to reconstruct accurate and extended phage genomes from MDA-treated virome samples of the human gut, and potentially from other environmental virome samples.

scholarly journals Long-read assemblies reveal structural diversity in genomes of organelles - an example with Acacia pycnantha

2020 ◽  
Author(s):  
Anna E. Syme ◽  
Todd G.B. McLay ◽  
Frank Udovicic ◽  
David J. Cantrill ◽  
Daniel J. Murphy
Keyword(s):  
Mitochondrial Genome ◽  
Chloroplast Genome ◽  
De Novo ◽  
Genomic Structure ◽  
Structural Diversity ◽  
Mitochondrial Genomes ◽  
Long Reads ◽  
Organelle Genomes ◽  
Long Read ◽  
Assembly Algorithms

AbstractAlthough organelle genomes are typically represented as single, static, circular molecules, there is evidence that the chloroplast genome exists in two structural haplotypes and that the mitochondrial genome can display multiple circular, linear or branching forms. We sequenced and assembled chloroplast and mitochondrial genomes of the Golden Wattle, Acacia pycnantha, using long reads, iterative baiting to extract organelle-only reads, and several assembly algorithms to explore genomic structure. Using a de novo assembly approach agnostic to previous hypotheses about structure, we found different assemblies revealed contrasting arrangements of genomic segments; a hypothesis supported by mapped reads spanning alternate paths.

scholarly journals Fast-SG: An alignment-free algorithm for hybrid assembly

2017 ◽  
Author(s):  
Alex Di Genova ◽  
Gonzalo A. Ruz ◽  
Marie-France Sagot ◽  
Alejandro Maass
Keyword(s):  
De Novo ◽  
Reference Level ◽  
Hybrid Assembly ◽  
Short Read ◽  
Sequencing Technologies ◽  
Alignment Free ◽  
Long Reads ◽  
Long Read ◽  
Definition Of ◽  
Large Genomes

ABSTRACTLong read sequencing technologies are the ultimate solution for genome repeats, allowing near reference level reconstructions of large genomes. However, long read de novo assembly pipelines are computationally intense and require a considerable amount of coverage, thereby hindering their broad application to the assembly of large genomes. Alternatively, hybrid assembly methods which combine short and long read sequencing technologies can reduce the time and cost required to produce de novo assemblies of large genomes. In this paper, we propose a new method, called FAST-SG, which uses a new ultra-fast alignment-free algorithm specifically designed for constructing a scaffolding graph using light-weight data structures. FAST-SG can construct the graph from either short or long reads. This allows the reuse of efficient algorithms designed for short read data and permits the definition of novel modular hybrid assembly pipelines. Using comprehensive standard datasets and benchmarks, we show how FAST-SG outperforms the state-of-the-art short read aligners when building the scaffolding graph, and can be used to extract linking information from either raw or error-corrected long reads. We also show how a hybrid assembly approach using FAST-SG with shallow long read coverage (5X) and moderate computational resources can produce long-range and accurate reconstructions of the genomes of Arabidopsis thaliana (Ler-0) and human (NA12878).

scholarly journals De novo genome assembly of the olive fruit fly (Bactrocera oleae) developed through a combination of linked-reads and long-read technologies

2018 ◽  
Author(s):  
Haig Djambazian ◽  
Anthony Bayega ◽  
Konstantina T. Tsoumani ◽  
Efthimia Sagri ◽  
Maria-Eleni Gregoriou ◽  
...  
Keyword(s):  
Y Chromosome ◽  
De Novo ◽  
Fruit Fly ◽  
Bactrocera Oleae ◽  
Olive Fruit Fly ◽  
Olive Fruit ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Oxford Nanopore Technologies

AbstractLong-read sequencing has greatly contributed to the generation of high quality assemblies, albeit at a high cost. It is also not always clear how to combine sequencing platforms. We sequenced the genome of the olive fruit fly (Bactrocera oleae), the most important pest in the olive fruits agribusiness industry, using Illumina short-reads, mate-pairs, 10x Genomics linked-reads, Pacific Biosciences (PacBio), and Oxford Nanopore Technologies (ONT). The 10x linked-reads assembly gave the most contiguous assembly with an N50 of 2.16 Mb. Scaffolding the linked-reads assembly using long-reads from ONT gave a more contiguous assembly with scaffold N50 of 4.59 Mb. We also present the most extensive transcriptome datasets of the olive fly derived from different tissues and stages of development. Finally, we used the Chromosome Quotient method to identify Y-chromosome scaffolds and show that the long-reads based assembly generates very highly contiguous Y-chromosome assembly.JR is a member of the MinION Access Program (MAP) and has received free-of-charge flow cells and sequencing kits from Oxford Nanopore Technologies for other projects. JR has had no other financial support from ONT.AB has received re-imbursement for travel costs associated with attending Nanopore Community meeting 2018, a meeting organized my Oxford Nanopore Technologies.

scholarly journals The evaluation of RNA-Seq de novo assembly by PacBio long read sequencing

2019 ◽  
Author(s):  
Yifan Yang ◽  
Michael Gribskov
Keyword(s):  
Real Time ◽  
De Novo ◽  
Critical Issue ◽  
Evaluation Methods ◽  
Model Organisms ◽  
Rna Seq ◽  
Long Reads ◽  
Long Read ◽  
Set Up ◽  
Downstream Analysis

AbstractRNA-Seq de novo assembly is an important method to generate transcriptomes for non-model organisms before any downstream analysis. Given many great de novo assembly methods developed by now, one critical issue is that there is no consensus on the evaluation of de novo assembly methods yet. Therefore, to set up a benchmark for evaluating the quality of de novo assemblies is very critical. Addressing this challenge will help us deepen the insights on the properties of different de novo assemblers and their evaluation methods, and provide hints on choosing the best assembly sets as transcriptomes of non-model organisms for the further functional analysis. In this article, we generate a “real time” transcriptome using PacBio long reads as a benchmark for evaluating five de novo assemblers and two model-based de novo assembly evaluation methods. By comparing the de novo assmblies generated by RNA-Seq short reads with the “real time” transcriptome from the same biological sample, we find that Trinity is best at the completeness by generating more assemblies than the alternative assemblers, but less continuous and having more misassemblies; Oases is best at the continuity and specificity, but less complete; The performance of SOAPdenovo-Trans, Trans-AByss and IDBA-Tran are in between of five assemblers. For evaluation methods, DETONATE leverages multiple aspects of the assembly set and ranks the assembly set with an average performance as the best, meanwhile the contig score can serve as a good metric to select assemblies with high completeness, specificity, continuity but not sensitive to misassemblies; TransRate contig score is useful for removing misassemblies, yet often the assemblies in the optimal set is too few to be used as a transcriptome.

scholarly journals RADAR-Base: Open Source Mobile Health Platform for Collecting, Monitoring, and Analyzing Data Using Sensors, Wearables, and Mobile Devices

10.2196/11734 ◽  
2019 ◽  
Vol 7 (8) ◽  
pp. e11734 ◽  
Author(s):  
Yatharth Ranjan ◽  
Zulqarnain Rashid ◽  
Callum Stewart ◽  
Pauline Conde ◽  
Mark Begale ◽  
...  
Keyword(s):  
High Resolution ◽  
Data Collection ◽  
Open Source ◽  
Mobile Health ◽  
Large Scale ◽  
Streaming Data ◽  
Security And Privacy ◽  
The European Union ◽  
Wide Range ◽  
Remote Assessment

Background With a wide range of use cases in both research and clinical domains, collecting continuous mobile health (mHealth) streaming data from multiple sources in a secure, highly scalable, and extensible platform is of high interest to the open source mHealth community. The European Union Innovative Medicines Initiative Remote Assessment of Disease and Relapse-Central Nervous System (RADAR-CNS) program is an exemplary project with the requirements to support the collection of high-resolution data at scale; as such, the Remote Assessment of Disease and Relapse (RADAR)-base platform is designed to meet these needs and additionally facilitate a new generation of mHealth projects in this nascent field. Objective Wide-bandwidth networks, smartphone penetrance, and wearable sensors offer new possibilities for collecting near-real-time high-resolution datasets from large numbers of participants. The aim of this study was to build a platform that would cater for large-scale data collection for remote monitoring initiatives. Key criteria are around scalability, extensibility, security, and privacy. Methods RADAR-base is developed as a modular application; the backend is built on a backbone of the highly successful Confluent/Apache Kafka framework for streaming data. To facilitate scaling and ease of deployment, we use Docker containers to package the components of the platform. RADAR-base provides 2 main mobile apps for data collection, a Passive App and an Active App. Other third-Party Apps and sensors are easily integrated into the platform. Management user interfaces to support data collection and enrolment are also provided. Results General principles of the platform components and design of RADAR-base are presented here, with examples of the types of data currently being collected from devices used in RADAR-CNS projects: Multiple Sclerosis, Epilepsy, and Depression cohorts. Conclusions RADAR-base is a fully functional, remote data collection platform built around Confluent/Apache Kafka and provides off-the-shelf components for projects interested in collecting mHealth datasets at scale.

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