Nejvíce citovaný článek - PubMed ID 28447635
A chromosome conformation capture ordered sequence of the barley genome
A pan-transcriptome describes the transcriptional and post-transcriptional consequences of genome diversity from multiple individuals within a species. We developed a barley pan-transcriptome using 20 inbred genotypes representing domesticated barley diversity by generating and analyzing short- and long-read RNA-sequencing datasets from multiple tissues. To overcome single reference bias in transcript quantification, we constructed genotype-specific reference transcript datasets (RTDs) and integrated these into a linear pan-genome framework to create a pan-RTD, allowing transcript categorization as core, shell or cloud. Focusing on the core (expressed in all genotypes), we observed significant transcript abundance variation among tissues and between genotypes driven partly by RNA processing, gene copy number, structural rearrangements and conservation of promotor motifs. Network analyses revealed conserved co-expression module::tissue correlations and frequent functional diversification. To complement the pan-transcriptome, we constructed a comprehensive cultivar (cv.) Morex gene-expression atlas and illustrate how these combined datasets can be used to guide biological inquiry.
Precise localization and dissection of gene promoters are key to understanding transcriptional gene regulation and to successful bioengineering applications. The core RNA polymerase II initiation machinery is highly conserved among eukaryotes, leading to a general expectation of equivalent underlying mechanisms. Still, less is known about promoters in the plant kingdom. In this study, we employed cap analysis of gene expression (CAGE) at three embryonic developmental stages in barley to accurately map, annotate, and quantify transcription initiation events. Unsupervised discovery of de novo sequence clusters grouped promoters based on characteristic initiator and position-specific core-promoter motifs. This grouping was complemented by the annotation of transcription factor binding site (TFBS) motifs. Integration with genome-wide epigenomic data sets and gene ontology (GO) enrichment analysis further delineated the chromatin environments and functional roles of genes associated with distinct promoter categories. The TATA-box presence governs all features explored, supporting the general model of two separate genomic regulatory environments. We describe the extent and implications of alternative transcription initiation events, including those that are specific to developmental stages, which can affect the protein sequence or the presence of regions that regulate translation. The generated promoterome dataset provides a valuable genomic resource for enhancing the functional annotation of the barley genome. It also offers insights into the transcriptional regulation of individual genes and presents opportunities for the informed manipulation of promoter architecture, with the aim of enhancing traits of agronomic importance.
- Klíčová slova
- Cap Analysis of Gene Expression, Core promoter, Hordeum vulgare, Initiator, Morex, TOR-signaling, Transcription regulation,
- Publikační typ
- časopisecké články MeSH
Cultivated and wild species of the genus rye (Secale) are important but underexploited gene sources for increasing the genetic diversity of bread wheat. Gene transfer is possible via bridge genetic materials derived from intergeneric hybrids. During this process, it is essential to precisely identify the rye chromatin in the wheat genetic background. In the present study, backcross generation BC2F8 from a cross between Triticum aestivum (Mv9kr1) and S. cereanum ('Kriszta,' a cultivar from the artificial hybrid of S. cereale and S. strictum) was screened using in-situ hybridization (GISH and FISH) and analyzed by DArTseq genotyping in order to select potentially agronomically useful genotypes for prebreeding purposes. Of the 329,267 high-quality short sequence reads generated, 27,822 SilicoDArT and 8,842 SNP markers specific to S. cereanum 1R-7R chromosomes were identified. Heatmaps of the marker densities along the 'Lo7' rye reference pseudomolecules revealed subtle differences between the FISH- and DArTseq-based results. This study demonstrates that the "exotic" rye chromatin of S. cereanum introgressed into wheat can be reliably identified by high-throughput DArTseq genotyping. The Mv9kr1-'Kriszta' addition and translocation lines presented here may serve as valuable prebreeding genetic materials for the development of stress-tolerant or disease-resistant wheat varieties.
- Klíčová slova
- DArTseq markers, Secale cereanum, Triticum aestivum, chromosome rearrangements, genotyping, heatmap, introgression lines,
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Roots play an important role during plant growth and development, ensuring water and nutrient uptake. Understanding the mechanisms regulating their initiation and development opens doors towards root system architecture engineering. RESULTS: Here, we investigated by RNA-seq analysis the changes in gene expression in the barley stem base of 1 day-after-germination (DAG) and 10DAG seedlings when crown roots are formed. We identified 2,333 genes whose expression was lower in the stem base of 10DAG seedlings compared to 1DAG seedlings. Those genes were mostly related to basal cellular activity such as cell cycle organization, protein biosynthesis, chromatin organization, cytoskeleton organization or nucleotide metabolism. In opposite, 2,932 genes showed up-regulation in the stem base of 10DAG seedlings compared to 1DAG seedlings, and their function was related to phytohormone action, solute transport, redox homeostasis, protein modification, secondary metabolism. Our results highlighted genes that are likely involved in the different steps of crown root formation from initiation to primordia differentiation and emergence, and revealed the activation of different hormonal pathways during this process. CONCLUSIONS: This whole transcriptomic study is the first study aiming at understanding the molecular mechanisms controlling crown root development in barley. The results shed light on crown root emergence that is likely associated with a strong cell wall modification, death of the cells covering the crown root primordium, and the production of defense molecules that might prevent pathogen infection at the site of root emergence.
- Klíčová slova
- Barley (Hordeum vulgare L.), Crown roots, Emergence, Transcriptome,
- MeSH
- ječmen (rod) * genetika růst a vývoj metabolismus MeSH
- kořeny rostlin * růst a vývoj genetika metabolismus MeSH
- regulace genové exprese u rostlin * MeSH
- rostlinné geny MeSH
- semenáček růst a vývoj genetika MeSH
- stanovení celkové genové exprese MeSH
- transkriptom MeSH
- Publikační typ
- časopisecké články MeSH
Despite more than a century of intensive study of mitotic chromosomes, their three-dimensional organization remains enigmatic. The last decade established Hi-C as a method of choice for study of spatial genome-wide interactions. Although its utilization has been focused mainly on studying genomic interactions in interphase nuclei, the method can be also successfully applied to study 3D architecture and genome folding in mitotic chromosomes. However, obtaining sufficient number of mitotic chromosomes as an input material and effective coupling with Hi-C method is challenging in plant species. An elegant way to overcome hindrances with obtaining a pure mitotic chromosome fraction is their isolation via flow cytometric sorting. This chapter presents a protocol describing plant sample preparation for chromosome conformation studies, for flow-sorting of plant mitotic metaphase chromosomes and for the Hi-C procedure.
- Klíčová slova
- 3D architecture, Chromatin interaction, Flow sorting, Hi-C, Plant chromosomes,
- MeSH
- buněčné jádro genetika MeSH
- chromatin * genetika MeSH
- chromozomy * genetika MeSH
- genomika metody MeSH
- molekulární konformace MeSH
- rostliny genetika MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- chromatin * MeSH
Optical mapping-a technique that visualizes short sequence motives along DNA molecules of hundred kilobases to megabase in size-has found an important place in genome research. It is widely used to facilitate genome sequence assemblies and analyses of genome structural variations. Application of the technique is conditional on availability of highly pure ultra-long high-molecular-weight DNA (uHMW DNA), which is challenging to achieve in plants due to the presence of the cell wall, chloroplasts, and secondary metabolites, just as a high content of polysaccharides and DNA nucleases in some species. These obstacles can be overcome by employment of flow cytometry, enabling a fast and highly efficient purification of cell nuclei or metaphase chromosomes, which are afterward embedded in agarose plugs and used to isolate the uHMW DNA in situ. Here, we provide a detailed protocol for the flow sorting-assisted uHMW DNA preparation that has been successfully used to construct whole-genome as well as chromosomal optical maps for 20 plant species from several plant families.
- Klíčová slova
- Bionano genome map, Chromosomes, Flow cytometry and sorting, HMW DNA preparation, Nuclei, Optical mapping, ultralong high-molecular-weight DNA,
- MeSH
- chromozomy rostlin * genetika MeSH
- genom rostlinný MeSH
- průtoková cytometrie metody MeSH
- restrikční mapování MeSH
- rostliny * genetika MeSH
- sekvenční analýza DNA metody MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Even though Sugars Will Eventually be Exported Transporters (SWEETs) have been found in every sequenced plant genome, a comprehensive understanding of their functionality is lacking. In this study, we focused on the SWEET family of barley (Hordeum vulgare). A radiotracer assay revealed that expressing HvSWEET11b in African clawed frog (Xenopus laevis) oocytes facilitated the bidirectional transfer of not only just sucrose and glucose, but also cytokinin. Barley plants harboring a loss-of-function mutation of HvSWEET11b could not set viable grains, while the distribution of sucrose and cytokinin was altered in developing grains of plants in which the gene was knocked down. Sucrose allocation within transgenic grains was disrupted, which is consistent with the changes to the cytokinin gradient across grains, as visualized by magnetic resonance imaging and Fourier transform infrared spectroscopy microimaging. Decreasing HvSWEET11b expression in developing grains reduced overall grain size, sink strength, the number of endopolyploid endosperm cells, and the contents of starch and protein. The control exerted by HvSWEET11b over sugars and cytokinins likely predetermines their synergy, resulting in adjustments to the grain's biochemistry and transcriptome.
- MeSH
- cukry metabolismus MeSH
- cytokininy * metabolismus MeSH
- ječmen (rod) * genetika metabolismus MeSH
- rostlinné proteiny genetika metabolismus MeSH
- sacharosa metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- cukry MeSH
- cytokininy * MeSH
- rostlinné proteiny MeSH
- sacharosa MeSH
Chromatids of mitotic chromosomes were suggested to coil into a helix in early cytological studies and this assumption was recently supported by chromosome conformation capture (3C) sequencing. Still, direct differential visualization of a condensed chromatin fibre confirming the helical model was lacking. Here, we combined Hi-C analysis of purified metaphase chromosomes, biopolymer modelling and spatial structured illumination microscopy of large fluorescently labeled chromosome segments to reveal the chromonema - a helically-wound, 400 nm thick chromatin thread forming barley mitotic chromatids. Chromatin from adjacent turns of the helix intermingles due to the stochastic positioning of chromatin loops inside the chromonema. Helical turn size varies along chromosome length, correlating with chromatin density. Constraints on the observable dimensions of sister chromatid exchanges further supports the helical chromonema model.
Increasing the proportion of locally produced plant protein in currently meat-rich diets could substantially reduce greenhouse gas emissions and loss of biodiversity1. However, plant protein production is hampered by the lack of a cool-season legume equivalent to soybean in agronomic value2. Faba bean (Vicia faba L.) has a high yield potential and is well suited for cultivation in temperate regions, but genomic resources are scarce. Here, we report a high-quality chromosome-scale assembly of the faba bean genome and show that it has expanded to a massive 13 Gb in size through an imbalance between the rates of amplification and elimination of retrotransposons and satellite repeats. Genes and recombination events are evenly dispersed across chromosomes and the gene space is remarkably compact considering the genome size, although with substantial copy number variation driven by tandem duplication. Demonstrating practical application of the genome sequence, we develop a targeted genotyping assay and use high-resolution genome-wide association analysis to dissect the genetic basis of seed size and hilum colour. The resources presented constitute a genomics-based breeding platform for faba bean, enabling breeders and geneticists to accelerate the improvement of sustainable protein production across the Mediterranean, subtropical and northern temperate agroecological zones.
- MeSH
- amplifikace genu genetika MeSH
- celogenomová asociační studie MeSH
- chromozomy rostlin genetika MeSH
- diploidie * MeSH
- genetická variace * genetika MeSH
- genom rostlinný * genetika MeSH
- genomika * MeSH
- rekombinace genetická MeSH
- retroelementy genetika MeSH
- rostlinné geny genetika MeSH
- rostlinné proteiny * genetika metabolismus MeSH
- satelitní DNA genetika MeSH
- semena rostlinná anatomie a histologie genetika MeSH
- šlechtění rostlin * metody MeSH
- variabilita počtu kopií segmentů DNA genetika MeSH
- Vicia faba * anatomie a histologie genetika metabolismus MeSH
- zemědělské plodiny * genetika metabolismus MeSH
- zeměpis MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- retroelementy MeSH
- rostlinné proteiny * MeSH
- satelitní DNA MeSH
Barley is the fifth most important food crop in Ethiopia. The genetic relationship and population structure studies of barley are limited to gene bank collections. Therefore, this study fills a gap by investigating the selection, consumption, economic value, genetic diversity, and population structure of farm-collected barley from the Gumer district of the Gurage Zone, which has received little attention. The information on the use of barley in the study area was collected using semi-structured interviews and questionnaires. 124 households of 11 kebeles, the smallest community unit, were interviewed. Barley landraces collected were compared with those collected from Japan, the United States (USA), and other Ethiopian locations. Illumina iSelect (50K genotyping platform) was used to identify single nucleotide polymorphisms (SNP) (20,367). Thirty landraces were found in Gumer. Burdaenadenber had the highest on-farm Shannon index estimate (2.0), followed by Aselecha (1.97) and Enjefo (1.95). Aselecha and Fetazer had the highest (44%) and the lowest (29%) richness values, respectively. High and low Simpson index values were found in Aselecha (84%) and Wulbaragenateretero (79%), respectively. The neighbor-joining tree revealed that Gumer landraces formed a separate subcluster with a common ancestral node; a sister subcluster contained barley landraces from Japan. According to the population structure analysis, barley landraces from Gumer differed from Japan and the United States. The principal component analysis revealed that US barley was the most distant group from Gumer barley. The markers' allele frequencies ranged from 0.10 to 0.50, with an average value of 0.28. The mean values of Nei's gene diversity (0.38) and the polymorphic information content (0.30) indicated the presence of high genetic diversity in the samples. The clustering of accessions was not based on geographic origin. Significant genetic diversity calls for additional research and analysis of local barley diversity because the selection and use of barley in Ethiopia would have been affected by the preference of ethnic groups.
- MeSH
- frekvence genu MeSH
- genetická variace * MeSH
- ječmen (rod) * genetika MeSH
- šlechtění rostlin MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Etiopie MeSH
