Sugarcane, the world's most harvested crop by tonnage, has shaped global history, trade and geopolitics, and is currently responsible for 80% of sugar production worldwide1. While traditional sugarcane breeding methods have effectively generated cultivars adapted to new environments and pathogens, sugar yield improvements have recently plateaued2. The cessation of yield gains may be due to limited genetic diversity within breeding populations, long breeding cycles and the complexity of its genome, the latter preventing breeders from taking advantage of the recent explosion of whole-genome sequencing that has benefited many other crops. Thus, modern sugarcane hybrids are the last remaining major crop without a reference-quality genome. Here we take a major step towards advancing sugarcane biotechnology by generating a polyploid reference genome for R570, a typical modern cultivar derived from interspecific hybridization between the domesticated species (Saccharum officinarum) and the wild species (Saccharum spontaneum). In contrast to the existing single haplotype ('monoploid') representation of R570, our 8.7 billion base assembly contains a complete representation of unique DNA sequences across the approximately 12 chromosome copies in this polyploid genome. Using this highly contiguous genome assembly, we filled a previously unsized gap within an R570 physical genetic map to describe the likely causal genes underlying the single-copy Bru1 brown rust resistance locus. This polyploid genome assembly with fine-grain descriptions of genome architecture and molecular targets for biotechnology will help accelerate molecular and transgenic breeding and adaptation of sugarcane to future environmental conditions.
- MeSH
- biotechnologie MeSH
- chromozomy rostlin genetika MeSH
- DNA rostlinná genetika MeSH
- genom rostlinný * genetika MeSH
- haplotypy genetika MeSH
- hybridizace genetická genetika MeSH
- polyploidie * MeSH
- referenční standardy MeSH
- Saccharum * klasifikace genetika MeSH
- šlechtění rostlin MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
- Názvy látek
- DNA rostlinná MeSH
This paper presents the latest update to the Plant rDNA database (Release 4.0), a valuable resource for researchers in the field of plant cytogenetics. The database provides information on the number, position, and arrangement of ribosomal DNA loci in plants, including angiosperms, gymnosperms, bryophytes, and pteridophytes. The new release includes new data for 820 species coming from additional 173 papers. In the updated version of the Plant rDNA database, 4948 entries comprising 2760 organisms can be found. A brief guide on how to navigate the database and obtain the desired information is also provided. The regular updating of the database is important for ensuring the information it contains is accurate, up-to-date, and useful for the research community. The Plant rDNA database continues to be beneficial for phylogenetic and cytogenetic studies in a wide range of taxa including angiosperms, gymnosperms, and early diverging groups, such as bryophytes and lycophytes.
- Klíčová slova
- Chromosome, Comparative cytogenetics, Data mining, In situ hybridization, Karyotype, L-type arrangement, Ribosomal DNA, S-type arrangement,
- MeSH
- cytogenetické vyšetření MeSH
- DNA rostlinná genetika MeSH
- fylogeneze MeSH
- informační zdroje * MeSH
- Magnoliopsida * MeSH
- ribozomální DNA genetika MeSH
- ribozomy MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA rostlinná MeSH
- ribozomální DNA MeSH
Flow cytometry has emerged as a uniquely flexible, accurate, and widely applicable technology for the analysis of plant cells. One of its most important applications centers on the measurement of nuclear DNA contents. This chapter describes the essential features of this measurement, outlining the overall methods and strategies, but going on to provide a wealth of technical details to ensure the most accurate and reproducible results. The chapter is aimed to be equally accessible to experienced plant cytometrists as well as those newly entering the field. Besides providing a step-by-step guide for estimating genome sizes and DNA-ploidy levels from fresh tissues, special attention is paid to the use of seeds and desiccated tissues for such purposes. Methodological aspects regarding field sampling, transport, and storage of plant material are also given in detail. Finally, troubleshooting information for the most common problems that may arise during the application of these methods is provided.
- Klíčová slova
- Best practices, DAPI, DNA-ploidy level, Desiccated tissues, Flow cytometry, Genome size, Plant nuclei isolation, Plant tissues, Propidium iodide, Seeds,
- MeSH
- buněčné jádro * genetika chemie MeSH
- délka genomu MeSH
- DNA rostlinná genetika analýza MeSH
- genom rostlinný MeSH
- ploidie MeSH
- průtoková cytometrie metody MeSH
- rostliny * genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA rostlinná MeSH
BACKGROUND AND AIMS: A targeted enrichment NGS approach was used to construct the phylogeny of Amomum Roxb. (Zingiberaceae). Phylogenies based on hundreds of nuclear genes, the whole plastome and the rDNA cistron were compared with an ITS-based phylogeny. Trends in genome size (GS) evolution were examined, chromosomes were counted and the geographical distribution of phylogenetic lineages was evaluated. METHODS: In total, 92 accessions of 54 species were analysed. ITS was obtained for 79 accessions, 37 accessions were processed with Hyb-Seq and sequences from 449 nuclear genes, the whole cpDNA, and the rDNA cistron were analysed using concatenation, coalescence and supertree approaches. The evolution of absolute GS was analysed in a phylogenetic and geographical context. The chromosome numbers of 12 accessions were counted. KEY RESULTS: Four groups were recognised in all datasets though their mutual relationships differ among datasets. While group A (A. subulatum and A. petaloideum) is basal to the remaining groups in the nuclear gene phylogeny, in the cpDNA topology it is sister to group B (A. repoeense and related species) and, in the ITS topology, it is sister to group D (the Elettariopsis lineage). The former Elettariopsis makes a monophyletic group. There is an increasing trend in GS during evolution. The largest GS values were found in group D in two tetraploid taxa, A. cinnamomeum and A. aff. biphyllum (both 2n = 96 chromosomes). The rest varied in GS (2C = 3.54-8.78 pg) with a constant chromosome number 2n = 48. There is a weak connection between phylogeny, GS and geography in Amomum. CONCLUSIONS: Amomum consists of four groups, and the former Elettariopsis is monophyletic. Species in this group have the largest GS. Two polyploids were found and GS greatly varied in the rest of Amomum.
- Klíčová slova
- Chromosome counts, Elettariopsis, Hyb-Seq, ITS, Polyploidy, cpDNA,
- MeSH
- Amomum * genetika MeSH
- délka genomu MeSH
- DNA chloroplastová MeSH
- DNA rostlinná genetika MeSH
- fylogeneze MeSH
- genom rostlinný MeSH
- ribozomální DNA genetika MeSH
- zázvorníkovité * genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA chloroplastová MeSH
- DNA rostlinná MeSH
- ribozomální DNA MeSH
The estimation of nuclear DNA content has been by far the most popular application of flow cytometry in plants. Because flow cytometry measures relative fluorescence intensities of nuclei stained by a DNA fluorochrome, ploidy determination, and estimation of the nuclear DNA content in absolute units both require comparison to a reference standard of known DNA content. This implies that the quality of the results obtained depends on the standard selection and use. Internal standardization, when the nuclei of an unknown sample and the reference standard are isolated, stained, and measured simultaneously, is mandatory for precise measurements. As DNA peaks representing G1 /G0 nuclei of the sample and standard appear on the same histogram of fluorescence intensity, the quotient of their position on the fluorescence intensity axis provides the quotient of DNA amounts. For the estimation of DNA amounts in absolute units, a number of well-established standards are now available to cover the range of known plant genome sizes. Since there are different standards in use, the standard and the genome size assigned to it has always to be reported. When none of the established standards fits, the introduction of a new standard species is needed. For this purpose, the regression line approach or simultaneous analysis of the candidate standard with several established standards should be prioritized. Moreover, the newly selected standard organism has to fulfill a number of requirements: it should be easy to identify and maintain, taxonomically unambiguous, globally available, with known genome size stability, lacking problematic metabolites, suitable for isolation of sufficient amounts of nuclei, and enabling measurements with low coefficients of variation of DNA peaks, hence suitable for the preparation of high quality samples.
- Klíčová slova
- C-value, GC content, best practices, flow cytometry, genome size, plant sciences, plant standard species, standardization,
- MeSH
- DNA rostlinná genetika MeSH
- genom rostlinný * MeSH
- ploidie * MeSH
- průtoková cytometrie metody MeSH
- referenční standardy MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Názvy látek
- DNA rostlinná MeSH
In theory, any plant tissue providing intact nuclei in sufficient quantity is suitable for nuclear DNA content estimation using flow cytometry (FCM). While this certainly opens a wide variety of possible applications of FCM, especially when compared to classical karyological techniques restricted to tissues with active cell division, tissue selection and quality may directly affect the precision (and sometimes even reliability) of FCM measurements. It is usually convenient to first consider the goals of the study to either aim for the highest possible accuracy of estimates (e.g., for inferring genome size, detecting homoploid intraspecific genome size variation, aneuploidy, among others), or to decide that histograms of reasonable resolution provide sufficient information (e.g., ploidy level screening within a single model species). Here, a set of best practices guidelines for selecting the optimal plant tissue for FCM analysis, sampling of material, and material preservation and storage are provided. In addition, factors potentially compromising the quality of FCM estimates of nuclear DNA content and data interpretation are discussed.
- Klíčová slova
- Best practices, flow cytometry, nuclear DNA content, nuclear suspensions, plant material collection, plant material preservation, plant material storage, plant sciences, tissue selection,
- MeSH
- buněčné jádro * chemie genetika MeSH
- DNA nádorová analýza MeSH
- DNA rostlinná genetika MeSH
- ploidie * MeSH
- průtoková cytometrie metody MeSH
- reprodukovatelnost výsledků MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Názvy látek
- DNA nádorová MeSH
- DNA rostlinná MeSH
Flow cytometry (FCM) is currently the most widely-used method to establish nuclear DNA content in plants. Since simple, 1-3-parameter, flow cytometers, which are sufficient for most plant applications, are commercially available at a reasonable price, the number of laboratories equipped with these instruments, and consequently new FCM users, has greatly increased over the last decade. This paper meets an urgent need for comprehensive recommendations for best practices in FCM for different plant science applications. We discuss advantages and limitations of establishing plant ploidy, genome size, DNA base composition, cell cycle activity, and level of endoreduplication. Applications of such measurements in plant systematics, ecology, molecular biology research, reproduction biology, tissue cultures, plant breeding, and seed sciences are described. Advice is included on how to obtain accurate and reliable results, as well as how to manage troubleshooting that may occur during sample preparation, cytometric measurements, and data handling. Each section is followed by best practice recommendations; tips as to what specific information should be provided in FCM papers are also provided.
- Klíčová slova
- DNA base composition, DNA content, cell cycle, endoreduplication, flow cytometric seed screening, genome size, in vitro cultures, intraspecific variation, ploidy,
- MeSH
- délka genomu MeSH
- DNA rostlinná genetika MeSH
- genom rostlinný MeSH
- ploidie * MeSH
- průtoková cytometrie metody MeSH
- rostliny * genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Názvy látek
- DNA rostlinná MeSH
Whole-genome duplications (WGDs) and chromosome rearrangements (CRs) play the key role in driving the diversification and evolution of plant lineages. Although the direct link between WGDs and plant diversification is well documented, relatively few studies focus on the evolutionary significance of CRs. The cruciferous tribe Thlaspideae represents an ideal model system to address the role of large-scale chromosome alterations in genome evolution, as most Thlaspideae species share the same diploid chromosome number (2n = 2x = 14). Here we constructed the genome structure in 12 Thlaspideae species, including field pennycress (Thlaspi arvense) and garlic mustard (Alliaria petiolata). We detected and precisely characterized genus- and species-specific CRs, mostly pericentric inversions, as the main genome-diversifying drivers in the tribe. We reconstructed the structure of seven chromosomes of an ancestral Thlaspideae genome, identified evolutionary stable chromosomes versus chromosomes prone to CRs, estimated the rate of CRs, and uncovered an allohexaploid origin of garlic mustard from diploid taxa closely related to A. petiolata and Parlatoria cakiloidea. Furthermore, we performed detailed bioinformatic analysis of the Thlaspideae repeatomes, and identified repetitive elements applicable as unique species- and genus-specific barcodes and chromosome landmarks. This study deepens our general understanding of the evolutionary role of CRs, particularly pericentric inversions, in plant genome diversification, and provides a robust base for follow-up whole-genome sequencing efforts.
- Klíčová slova
- Brassicaceae, Thlaspideae, chromosome rearrangements, garlic mustard, genome evolution, pennycress, pericentric inversions, repetitive DNA,
- MeSH
- biologická evoluce MeSH
- Brassicaceae genetika MeSH
- chromozomální inverze MeSH
- chromozomy rostlin * MeSH
- diploidie MeSH
- DNA rostlinná genetika MeSH
- genom rostlinný * MeSH
- karyotyp MeSH
- repetitivní sekvence nukleových kyselin MeSH
- ribozomální DNA genetika MeSH
- Thlaspi genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA rostlinná MeSH
- ribozomální DNA MeSH
Kleisin NSE4 and circular form of SMC5/6 is indispensable for DSB repair and necessary for gene targeting but is not enough for recovery of cells from DNA damage in Physcomitrella. Structural maintenance of chromosomes (SMC) complexes are involved in cohesion, condensation and maintenance of genome stability. Based on the sensitivity of mutants to genotoxic stress the SMC5/6 complex is thought to play a prominent role in DNA stabilization during repair by tethering DNA at the site of lesion by a heteroduplex of SMC5 and SMC6 encircled with non-SMC components NSE1, NSE3 and kleisin NSE4. In this study, we tested how formation of the SMC5/6 circular structure affects mutant sensitivity to DNA damage, kinetics of DSB repair and gene targeting. In the moss Physcomitrella (Physcomitrium patens), SMC6 and NSE4 are essential single copy genes and this is why we used blocking of transcription to reveal their mutated phenotype. Even slight reduction of transcript levels by dCas9 binding was enough to obtain stable lines with severe DSB repair defects and specific bleomycin sensitivity. We show that survival after bleomycin or MMS treatment fully depends on active SMC6, whereas attenuation of NSE4 has little or negligible effect. We conclude that circularization of SMC5/6 provided by the kleisin NSE4 is indispensable for the DSB repair, nevertheless there are other functions associated with the SMC5/6 complex, which are critical to survive DNA damage.
- Klíčová slova
- Comet assay, DNA repair, Gene targeting, NSE4 kleisin, Physcomitrella, Physcomitrium patens, SMC5/6 complex, dCas9,
- MeSH
- DNA rostlinná genetika metabolismus MeSH
- dvouřetězcové zlomy DNA * MeSH
- fylogeneze MeSH
- genotyp MeSH
- mechy genetika metabolismus MeSH
- multiproteinové komplexy genetika metabolismus MeSH
- mutace MeSH
- oprava DNA * MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- poškození DNA * MeSH
- proteiny buněčného cyklu klasifikace genetika metabolismus MeSH
- regulace genové exprese u rostlin MeSH
- rostlinné proteiny klasifikace genetika metabolismus MeSH
- sekvence aminokyselin MeSH
- sekvenční homologie aminokyselin MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA rostlinná MeSH
- multiproteinové komplexy MeSH
- proteiny buněčného cyklu MeSH
- rostlinné proteiny MeSH
The aim of the study was to verify the hypothesis that a potential cause of the phytotoxicity of diclofenac (DCF, a non-steroidal anti-inflammatory drug) is an effect of cell cycle progression. This research was conducted using synchronous cultures of a model organism, green alga Chlamydomonas reinhardtii. The project examined DCF effects on selected parameters that characterize cell cycle progression, such as cell size, attainment of commitment points, DNA replication, number of nuclei formed during cells division and morphology of cells in consecutive stages of the cell cycle, together with the physiological and biochemical parameters of algae cells at different stages. We demonstrated that individual cell growth remained unaffected, whereas cell division was delayed in the DCF-treated groups grown in continuous light conditions, and the number of daughter cells from a single cell decreased. Thus, the cell cycle progression is a target affected by DCF, which has a similar anti-proliferative effect on mammalian cells.
- Klíčová slova
- Chlamydomonas reinhardtii, cell cycle, diclofenac, non-steroidal anti-inflammatory drug,
- MeSH
- antiflogistika nesteroidní toxicita MeSH
- buněčné dělení účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- Chlamydomonas reinhardtii účinky léků genetika růst a vývoj MeSH
- diklofenak toxicita MeSH
- DNA rostlinná biosyntéza genetika MeSH
- fotosyntéza účinky léků MeSH
- replikace DNA účinky léků MeSH
- velikost buňky účinky léků MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- antiflogistika nesteroidní MeSH
- diklofenak MeSH
- DNA rostlinná MeSH
