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Current Protocols in Microbiology Aug 2018Ploidy, the number of sets of homologous chromosomes in a cell, can alter cellular physiology, gene regulation, and the spectrum of acquired mutations. Advances in...
Ploidy, the number of sets of homologous chromosomes in a cell, can alter cellular physiology, gene regulation, and the spectrum of acquired mutations. Advances in single-cell flow cytometry have greatly improved the understanding of how genome size contributes to diverse biological processes including speciation, adaptation, pathogenesis, and tumorigenesis. For example, fungal pathogens can undergo whole genome duplications during infection of the human host and during acquisition of antifungal drug resistance. Quantification of ploidy is dramatically affected by the nucleic acid staining technique and the flow cytometry analysis of single cells. Ploidy in fungi is also impacted by samples that are heterogeneous for both ploidy and morphology, and control strains with known ploidy must be included in every flow cytometry experiment. To detect ploidy changes within fungal strains, the following protocol was developed to accurately and dependably interrogate single-cell ploidy. © 2018 by John Wiley & Sons, Inc.
Topics: Flow Cytometry; Fungi; Humans; Mycology; Mycoses; Ploidies; Software; Staining and Labeling
PubMed: 30028911
DOI: 10.1002/cpmc.58 -
Cytometry. Part a : the Journal of the... Sep 2022The estimation of nuclear DNA content has been by far the most popular application of flow cytometry in plants. Because flow cytometry measures relative fluorescence... (Review)
Review
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 G /G 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.
Topics: DNA, Plant; Flow Cytometry; Genome, Plant; Ploidies; Reference Standards
PubMed: 34405937
DOI: 10.1002/cyto.a.24495 -
Molecular Biology and Evolution Dec 2021Hybridization and genome duplication have played crucial roles in the evolution of many animal and plant taxa. The subgenomes of parental species undergo considerable...
Hybridization and genome duplication have played crucial roles in the evolution of many animal and plant taxa. The subgenomes of parental species undergo considerable changes in hybrids and polyploids, which often selectively eliminate segments of one subgenome. However, the mechanisms underlying these changes are not well understood, particularly when the hybridization is linked with asexual reproduction that opens up unexpected evolutionary pathways. To elucidate this problem, we compared published cytogenetic and RNAseq data with exome sequences of asexual diploid and polyploid hybrids between three fish species; Cobitis elongatoides, C. taenia, and C. tanaitica. Clonal genomes remained generally static at chromosome-scale levels but their heterozygosity gradually deteriorated at the level of individual genes owing to allelic deletions and conversions. Interestingly, the impact of both processes varies among animals and genomic regions depending on ploidy level and the properties of affected genes. Namely, polyploids were more tolerant to deletions than diploid asexuals where conversions prevailed, and genomic restructuring events accumulated preferentially in genes characterized by high transcription levels and GC-content, strong purifying selection and specific functions like interacting with intracellular membranes. Although hybrids were phenotypically more similar to C. taenia, we found that they preferentially retained C. elongatoides alleles. This demonstrates that favored subgenome is not necessarily the transcriptionally dominant one. This study demonstrated that subgenomes in asexual hybrids and polyploids evolve under a complex interplay of selection and several molecular mechanisms whose efficiency depends on the organism's ploidy level, as well as functional properties and parental ancestry of the genomic region.
Topics: Animals; Cypriniformes; Diploidy; Evolution, Molecular; Genome, Plant; Hybridization, Genetic; Loss of Heterozygosity; Polyploidy
PubMed: 34410426
DOI: 10.1093/molbev/msab249 -
EMBO Reports Dec 2020Polyploidy, a balanced amplification of the genome, is common in the liver. The function of hepatic polyploidy is not entirely clear, but growing evidence shows that...
Polyploidy, a balanced amplification of the genome, is common in the liver. The function of hepatic polyploidy is not entirely clear, but growing evidence shows that polyploidy can protect the liver from tumor formation. In this issue of EMBO Reports, Sladky and colleagues identify the PIDDosome as a polyploidy sensor that regulates liver cancer (Sladky et al, 2020b).
Topics: Hepatocytes; Humans; Liver; Liver Neoplasms; Ploidies; Polyploidy; Tumor Suppressor Protein p53
PubMed: 33237586
DOI: 10.15252/embr.202051922 -
Current Opinion in Plant Biology Jun 2021Interest in understanding hybrid seed failure (HSF) has mushroomed, both in terms of identifying underlying molecular processes and their evolutionary drivers. We review... (Review)
Review
Interest in understanding hybrid seed failure (HSF) has mushroomed, both in terms of identifying underlying molecular processes and their evolutionary drivers. We review phenotypic and molecular advances with a focus on the 'effective ploidy' concept, witnessing a recent revival after long obscurity. Endosperm misdevelopment has now been shown to underlie HSF in many inter-specific, homoploid crosses. The consistent asymmetries in seed size and developmental trajectories likely reflect parental divergence in key, dosage-sensitive processes. Transcriptomic and epigenomic studies reveal genome-wide, polarized expression perturbations and shifts in parental expression proportions, consistent with small-RNA imbalances between parental roles. Among-species differences in levels of parental conflict over resource allocation enjoy strong support in explaining why differences in effective ploidy may evolve.
Topics: Biological Evolution; Endosperm; Hybridization, Genetic; Ploidies; Reproductive Isolation; Seeds
PubMed: 33639340
DOI: 10.1016/j.pbi.2021.102015 -
Genes Jul 2023is the fastest-growing bacterium, with a doubling time of approximately 12-14 min. It has a high potential for basic research and biotechnological applications, e.g.,... (Review)
Review
is the fastest-growing bacterium, with a doubling time of approximately 12-14 min. It has a high potential for basic research and biotechnological applications, e.g., it can be used for the cell-free production of (labeled) heterologous proteins, for synthetic biological applications, and for the production of various compounds. However, the ploidy level in remains unknown. At nine time points throughout the growth curve, we analyzed the numbers of origins and termini of both chromosomes with qPCR and the relative abundances of all genomic sites with marker frequency analyses. During the lag phase until early exponential growth, the origin copy number and origin/terminus ratio of chromosome 1 increased severalfold, but the increase was lower for chromosome 2. This increase was paralleled by an increase in cell volume. During the exponential phase, the origin/terminus ratio and cell volume decreased again. This highly dynamic and fast regulation has not yet been described for any other species. In this study, the gene dosage increase in origin-adjacent genes during the lag phase is discussed together with the nonrandom distribution of genes on the chromosomes of . Taken together, the results of this study provide the first comprehensive overview of the chromosome dynamics in and will guide the optimization of molecular biological characterization and biotechnological applications.
Topics: DNA Copy Number Variations; Vibrio; Chromosomes; Ploidies
PubMed: 37510340
DOI: 10.3390/genes14071437 -
Journal of Visualized Experiments : JoVE Mar 2018Mechanisms that involve whole genome polyploidy play important roles in development and evolution; also, an abnormal generation of tetraploid cells has been associated...
Mechanisms that involve whole genome polyploidy play important roles in development and evolution; also, an abnormal generation of tetraploid cells has been associated with both the progression of cancer and the development of drug resistance. Until now, it has not been feasible to easily manipulate the ploidy of a multicellular animal without generating mostly sterile progeny. Presented here is a simple and rapid protocol for generating tetraploid Caenorhabditis elegans animals from any diploid strain. This method allows the user to create a bias in chromosome segregation during meiosis, ultimately increasing ploidy in C. elegans. This strategy relies on the transient reduction of expression of the rec-8 gene to generate diploid gametes. A rec-8 mutant produces diploid gametes that can potentially produce tetraploids upon fertilization. This tractable scheme has been used to generate tetraploid strains carrying mutations and chromosome rearrangements to gain insight into chromosomal dynamics and interactions during pairing and synapsis in meiosis. This method is efficient for generating stable tetraploid strains without genetic markers, can be applied to any diploid strain, and can be used to derive triploid C. elegans. This straightforward method is useful for investigating other fundamental biological questions relevant to genome instability, gene dosage, biological scaling, extracellular signaling, adaptation to stress, development of resistance to drugs, and mechanisms of speciation.
Topics: Animals; Caenorhabditis elegans; Male; Ploidies
PubMed: 29608173
DOI: 10.3791/57296 -
Cellular and Molecular Gastroenterology... 2020
Topics: Diploidy; Gene Expression Profiling; Hepatocytes; Humans; Ploidies; Polyploidy; Stem Cells; Transcriptome
PubMed: 31654613
DOI: 10.1016/j.jcmgh.2019.09.008 -
Bioinformatics (Oxford, England) Aug 2019CNValidator assesses the quality of somatic copy-number calls based on coherency of haplotypes across multiple samples from the same individual. It is applicable to any...
MOTIVATION
CNValidator assesses the quality of somatic copy-number calls based on coherency of haplotypes across multiple samples from the same individual. It is applicable to any copy-number calling algorithm, which makes calls independently for each sample. This test is useful in assessing the accuracy of copy-number calls, as well as choosing among alternative copy-number algorithms or tuning parameter values.
RESULTS
On a dataset of somatic samples from individuals with Barrett's Esophagus, CNValidator provided feedback on the correctness of sample ploidy calls and also detected data quality issues.
AVAILABILITY AND IMPLEMENTATION
CNValidator is available on GitHub at https://github.com/kuhnerlab/CNValidator.
SUPPLEMENTARY INFORMATION
Supplementary data are available at Bioinformatics online.
Topics: Algorithms; DNA Copy Number Variations; Haplotypes; Humans; Ploidies; Software
PubMed: 30541069
DOI: 10.1093/bioinformatics/bty1022 -
Annals of Botany Feb 2023Diploid and polyploid Urochloa (including Brachiaria, Panicum and Megathyrsus species) C4 tropical forage grasses originating from Africa are important for food security...
BACKGROUND AND AIMS
Diploid and polyploid Urochloa (including Brachiaria, Panicum and Megathyrsus species) C4 tropical forage grasses originating from Africa are important for food security and the environment, often being planted in marginal lands worldwide. We aimed to characterize the nature of their genomes, the repetitive DNA and the genome composition of polyploids, leading to a model of the evolutionary pathways within the group including many apomictic species.
METHODS
Some 362 forage grass accessions from international germplasm collections were studied, and ploidy was determined using an optimized flow cytometry method. Whole-genome survey sequencing and molecular cytogenetic analysis were used to identify chromosomes and genomes in Urochloa accessions belonging to the 'brizantha' and 'humidicola' agamic complexes and U. maxima.
KEY RESULTS
Genome structures are complex and variable, with multiple ploidies and genome compositions within the species, and no clear geographical patterns. Sequence analysis of nine diploid and polyploid accessions enabled identification of abundant genome-specific repetitive DNA motifs. In situ hybridization with a combination of repetitive DNA and genomic DNA probes identified evolutionary divergence and allowed us to discriminate the different genomes present in polyploids.
CONCLUSIONS
We suggest a new coherent nomenclature for the genomes present. We develop a model of evolution at the whole-genome level in diploid and polyploid accessions showing processes of grass evolution. We support the retention of narrow species concepts for Urochloa brizantha, U. decumbens and U. ruziziensis, and do not consider diploids and polyploids of single species as cytotypes. The results and model will be valuable in making rational choices of parents for new hybrids, assist in use of the germplasm for breeding and selection of Urochloa with improved sustainability and agronomic potential, and assist in measuring and conserving biodiversity in grasslands.
Topics: Poaceae; Brachiaria; Polyploidy; Ploidies; Genomics
PubMed: 34874999
DOI: 10.1093/aob/mcab147