De Novo Assembly of Complete Chloroplast Genomes from Non-model Species Based on a K-mer Frequency-Based Selection of Chloroplast Reads from Total DNA Sequences

Izan, Shairul; Esselink, Danny; Visser, Richard G. F.; Smulders, M.J.M.; Borm, Theo

doi:10.3389/fpls.2017.01271

Cited by 20 publications

(32 citation statements)

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“…First and foremost, it must be decided if full chloroplast genomes are necessary or if protein-coding gene space will be adequate. Acquiring full chloroplast genomes for each sample can add extra time to data generation and analysis; however, multiple assembly pipelines (e.g., ACRE [Wysocki et al, 2014], IOGA [Bakker et al, 2016], NOVOPlasty [Dierckxsens et al, 2017], Fast-Plast [https:// github.com/mrmckain/Fast-Plast], and a k-mer-based approach [Izan et al, 2017]) have been developed that are capable of assembling complete chloroplast genomes from short-read data. Complete chloroplast genomes can potentially provide more phylogenetic signal from intergenic regions for reconstructing relationships among closely related species (Carbonell-Caballero et al, 2015).…”

Section: Chloroplast Genomes Are Readily and Inexpensively Obtainedmentioning

confidence: 99%

Practical considerations for plant phylogenomics

et al. 2018

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The past decade has seen a major breakthrough in our ability to easily and inexpensively sequence genome‐scale data from diverse lineages. The development of high‐throughput sequencing and long‐read technologies has ushered in the era of phylogenomics, where hundreds to thousands of nuclear genes and whole organellar genomes are routinely used to reconstruct evolutionary relationships. As a result, understanding which options are best suited for a particular set of questions can be difficult, especially for those just starting in the field. Here, we review the most recent advances in plant phylogenomic methods and make recommendations for project‐dependent best practices and considerations. We focus on the costs and benefits of different approaches in regard to the information they provide researchers and the questions they can address. We also highlight unique challenges and opportunities in plant systems, such as polyploidy, reticulate evolution, and the use of herbarium materials, identifying optimal methodologies for each. Finally, we draw attention to lingering challenges in the field of plant phylogenomics, such as reusability of data sets, and look at some up‐and‐coming technologies that may help propel the field even further.

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Section: Chloroplast Genomes Are Readily and Inexpensively Obtainedmentioning

confidence: 99%

Practical considerations for plant phylogenomics

et al. 2018

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“…However, Geneious is closed source and GUI based, which is not in the scope of this study. There is also another publication describing a method for assembling chloroplasts [54]. However, the link to the software is not active anymore.…”

Section: Tool Selectionmentioning

confidence: 99%

A systematic comparison of chloroplast genome assembly tools

Freudenthal

Pfaff

Terhoeven

et al. 2019

Preprint

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Chloroplasts are photosynthetic organelles in plant cells and contain their own genomic information. That genome can be utilized in different scientific fields like phylogenetics or biotechnology. Thus, different assemblers have been developed specialized in chloroplast assemblies. Those assemblers often use the output of whole genome sequencing experiments as input. Such sequencing data usually contain the complete chloroplast genome information, even if the sequencing aims for the core genome. Different assembly tools have never been systematically compared. Here we present a benchmark of seven chloroplast assembly tools, capable to succeed in more than 60 % of real data sets. Our results show significant differences between the tested assemblers in terms of generating whole chloroplast genome sequences and computational requirements. Moreover, we suggest further development to improve user experience and success rate. In terms of reproducibility, we created docker images for each tested tool, which are available for the scientific community. Following the presented guidelines, users are able to analyze and screen data sets for chloroplast genomes using only standard computer infrastructure. Thus large scale screening for chloroplasts as hidden treasures within genomic sequencing data is feasible.

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“…The reason for that is that a mapping consensus will always adopt characteristics of the reference from which it was inferred. Structural rearrangements or additional transferred genes might easily be missed, especially when mapping short reads; the structure of repeat regions cannot be distinguished by mapping based approaches at all [86]. For the assembly of the globe artichoke (Cynara L.) cp genome with Illumina reads, [76] compared mapping and de novo approaches and found the results to be almost identical.…”

Section: Reference-based Assembly Using Nanopore Datamentioning

confidence: 99%

“…recognized an effect of too low or high coverage on assembly quality, stating that "there is a complex relationship between assembly accuracy and input read coverage when assembling chloroplast genomes with ONT data using Canu" [21]. [6] mention that de novo assembly performance could be improved by downsampling the input reads, and [86] stress that very high coverage may result in alternative assemblies which can lead to contig fragmentation. For the present study, preliminary de novo assembly tests (results not shown) with a reduced compared to the full dataset considerably decreased the number of contigs (approximately by half).…”

Section: Optimizing Nanopore De Novo and Reference-based Assemblymentioning

confidence: 99%

Can we use it? On the utility ofde novoand reference-based assembly of Nanopore data for plant plastome sequencing

Scheunert

Dorfner

Lingl

et al. 2019

Preprint

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The chloroplast genome harbors plenty of valuable information for phylogenetic research.Illumina short-read data is generally used for de novo assembly of whole plastomes. PacBio or Oxford Nanopore long reads are additionally employed in hybrid approaches to enable assembly across the highly similar inverted repeats of a chloroplast genome. Unlike for PacBio, plastome assemblies based solely on Nanopore reads are rarely found, due to their high error rate and non-random error profile. However, the actual quality decline connected to their use has never been quantified. Furthermore, no study has employed reference-based assembly using Nanopore reads, which is common with Illumina data. Using Leucanthemum Mill. as an example, we compared the sequence quality of seven plastome assemblies of the same species, using combinations of two sequencing platforms and three analysis pipelines.In addition, we assessed the factors which might influence Nanopore assembly quality during sequence generation and bioinformatic processing.The consensus sequence derived from de novo assembly of Nanopore data had a sequence identity of 99.59% compared to Illumina short-read de novo assembly. Most of the found errors comprise indels (81.5%), and a large majority of them is part of homopolymer regions.The quality of reference-based assembly is heavily dependent upon the choice of a closeenough reference. Using a reference with 0.83% sequence divergence from the studied species, mapping of Nanopore reads results in a consensus comparable to that from Nanopore de novo assembly, and of only slightly inferior quality compared to a referencebased assembly with Illumina data (0.49% and 0.26% divergence from Illumina de novo). For optimal assembly of Nanopore data, appropriate filtering of contaminants and chimeric sequences, as well as employing moderate read coverage, is essential.Based on these results, we conclude that Nanopore long reads are a suitable alternative to Illumina short reads in plastome phylogenomics. Only few errors remain in the finalized assembly, which can be easily masked in phylogenetic analyses without loss in analytical accuracy. The easily applicable and cost-effective technology might warrant more attention by researchers dealing with plant chloroplast genomes.

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De Novo Assembly of Complete Chloroplast Genomes from Non-model Species Based on a K-mer Frequency-Based Selection of Chloroplast Reads from Total DNA Sequences

Cited by 20 publications

References 50 publications

Practical considerations for plant phylogenomics

Practical considerations for plant phylogenomics

A systematic comparison of chloroplast genome assembly tools

Can we use it? On the utility ofde novoand reference-based assembly of Nanopore data for plant plastome sequencing

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