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

Istace, Benjamin; Belser, Caroline; Falentin, Cyril; Labadie, Karine; Boideau, Franz; Deniot, Gwenaëlle; Maillet, Loeiz; Cruaud, Corinne; Bertrand, Laurie; Chèvre, Anne‐Marie; Wincker, Patrick; Rousseau‐Gueutin, Mathieu; Aury, Jean‐Marc

doi:10.3390/biology10080732

Cited by 15 publications

(15 citation statements)

References 49 publications

(56 reference statements)

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“…First, we calculated the overall quality of the sequence using Merqury and Illumina reads. We obtained an average quality value (QV) of 32.8, a lower QV than that obtained with short-reads assemblies, but consistent with QV already reported for plant genomes sequenced by ONT[24]. Indeed, using Illumina reads and the CS RefSeq v2.1 assembly, Merqury computed a QV of 44.5 (Table 1).…”

Section: Resultssupporting

confidence: 85%

Long-read and chromosome-scale assembly of the hexaploid wheat genome achieves high resolution for research and breeding

Aury

Engelen

Istace

et al. 2021

Preprint

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The sequencing of the wheat (Triticum aestivum) genome has been a methodological challenge for many years due to its large size (15.5 Gb), repeat content, and hexaploidy. Many initiatives aiming at obtaining a reference genome of cultivar Chinese Spring have been launched in the past years and it was achieved in 2018 as the result of a huge effort to combine short-read whole genome sequencing with many other resources. Reference-quality genome assemblies were then produced for other accessions but the rapid evolution of sequencing technologies offers opportunities to reach high-quality standards at lower cost. Here, we report on an optimized procedure based on long-reads produced on the ONT (Oxford Nanopore Technology) PromethION device to assemble the genome of the French bread wheat cultivar Renan. We provide the most contiguous and complete chromosome-scale assembly of a bread wheat genome to date, a resource that will be valuable for the crop community and will facilitate the rapid selection of agronomically important traits. We also provide the methodological standards to generate high-quality assemblies of complex genomes.

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Section: Resultssupporting

confidence: 85%

Long-read and chromosome-scale assembly of the hexaploid wheat genome achieves high resolution for research and breeding

Aury

Engelen

Istace

et al. 2021

Preprint

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“…Darmor-bzh v10 (Rousseau-Gueutin et al, 2020) (Table S3). Centromeric regions were determined using the method described in Zhang et al (2018) and Istace et al (2021). Briefly, we retrieved the localization of the centromeric-specific repeat sequences using BLASTN (Altschul et al, 1990) and a cutoff of one-third of the maximal density identified on the chromosome.…”

Section: Darmor-bzh V10 Genomementioning

confidence: 99%

Epigenomic and structural events preclude recombination in Brassica napus

et al. 2022

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Meiotic recombination is a major evolutionary process generating genetic diversity at each generation in sexual organisms. However, this process is highly regulated, with the majority of crossovers lying in the distal chromosomal regions that harbor low DNA methylation levels. Even in these regions, some islands without recombination remain, for which we investigated the underlying causes.Genetic maps were established in two Brassica napus hybrids to detect the presence of such large nonrecombinant islands. The role played by DNA methylation and structural variations in this local absence of recombination was determined by performing bisulfite sequencing and whole genome comparisons. Inferred structural variations were validated using either optical mapping or oligo fluorescence in situ hybridization.Hypermethylated or inverted regions between Brassica genomes were associated with the absence of recombination. Pairwise comparisons of nine B. napus genome assemblies revealed that such inversions occur frequently and may contain key agronomic genes such as resistance to biotic stresses.We conclude that such islands without recombination can have different origins, such as DNA methylation or structural variations in B. napus. It is thus essential to take into account these features in breeding programs as they may hamper the efficient combination of favorable alleles in elite varieties.

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“…Long-read sequencing technologies revolutionized plant genomics both in terms of study frameworks and methodology [ 40 , 62 ]. Long reads enable researchers to assemble large plant genomes de novo with improved contiguity, compared to the shotgun sequencing approach [ 63 , 64 ]. However, the third-generation sequencing technologies have their own limitations.…”

Section: Discussionmentioning

confidence: 99%

“…Assembling a genome from ONT and PacBio reads simultaneously allowed researchers to delve deep into genome structure, track rearrangements, and detect more transposable elements [ 36 , 79 , 83 ]. Other options are the use of optical maps and Hi-C scaffolding, which successfully link contigs to chromosomes The received genomes become powerful sources of genetic information [ 62 , 64 , 84 ]. Therefore, the received L. usitatissimum genome can be upgraded to a chromosome scale and used in comprehensive structure analyses.…”

Section: Discussionmentioning

confidence: 99%

Isolating Linum usitatissimum L. Nuclear DNA Enabled Assembling High-Quality Genome

Dvorianinova

Bolsheva

Pushkova

et al. 2022

IJMS

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High-quality genome sequences help to elucidate the genetic basis of numerous biological processes and track species evolution. For flax (Linum usitatissimum L.)—a multifunctional crop, high-quality assemblies from Oxford Nanopore Technologies (ONT) data were unavailable, largely due to the difficulty of isolating pure high-molecular-weight DNA. This article proposes a scheme for gaining a contiguous L. usitatissimum assembly using Nanopore data. We developed a protocol for flax nuclei isolation with subsequent DNA extraction, which allows obtaining about 5 μg of pure high-molecular-weight DNA from 0.5 g of leaves. Such an amount of material can be collected even from a single plant and yields more than 30 Gb of ONT data in two MinION runs. We performed a comparative analysis of different genome assemblers and polishers on the gained data and obtained the final 447.1-Mb assembly of L. usitatissimum line 3896 genome using the Canu—Racon (two iterations)—Medaka combination. The genome comprised 1695 contigs and had an N50 of 6.2 Mb and a completeness of 93.8% of BUSCOs from eudicots_odb10. Our study highlights the impact of the chosen genome construction strategy on the resulting assembly parameters and its eligibility for future genomic studies.

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

Cited by 15 publications

References 49 publications

Long-read and chromosome-scale assembly of the hexaploid wheat genome achieves high resolution for research and breeding

Long-read and chromosome-scale assembly of the hexaploid wheat genome achieves high resolution for research and breeding

Epigenomic and structural events preclude recombination in Brassica napus

Isolating Linum usitatissimum L. Nuclear DNA Enabled Assembling High-Quality Genome

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