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1.
Gigascience ; 112022 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-35333302

RESUMO

BACKGROUND: Cassava (Manihot esculenta) is an important clonally propagated food crop in tropical and subtropical regions worldwide. Genetic gain by molecular breeding has been limited, partially because cassava is a highly heterozygous crop with a repetitive and difficult-to-assemble genome. FINDINGS: Here we demonstrate that Pacific Biosciences high-fidelity (HiFi) sequencing reads, in combination with the assembler hifiasm, produced genome assemblies at near complete haplotype resolution with higher continuity and accuracy compared to conventional long sequencing reads. We present 2 chromosome-scale haploid genomes phased with Hi-C technology for the diploid African cassava variety TME204. With consensus accuracy >QV46, contig N50 >18 Mb, BUSCO completeness of 99%, and 35k phased gene loci, it is the most accurate, continuous, complete, and haplotype-resolved cassava genome assembly so far. Ab initio gene prediction with RNA-seq data and Iso-Seq transcripts identified abundant novel gene loci, with enriched functionality related to chromatin organization, meristem development, and cell responses. During tissue development, differentially expressed transcripts of different haplotype origins were enriched for different functionality. In each tissue, 20-30% of transcripts showed allele-specific expression (ASE) differences. ASE bias was often tissue specific and inconsistent across different tissues. Direction-shifting was observed in <2% of the ASE transcripts. Despite high gene synteny, the HiFi genome assembly revealed extensive chromosome rearrangements and abundant intra-genomic and inter-genomic divergent sequences, with large structural variations mostly related to LTR retrotransposons. We use the reference-quality assemblies to build a cassava pan-genome and demonstrate its importance in representing the genetic diversity of cassava for downstream reference-guided omics analysis and breeding. CONCLUSIONS: The phased and annotated chromosome pairs allow a systematic view of the heterozygous diploid genome organization in cassava with improved accuracy, completeness, and haplotype resolution. They will be a valuable resource for cassava breeding and research. Our study may also provide insights into developing cost-effective and efficient strategies for resolving complex genomes with high resolution, accuracy, and continuity.


Assuntos
Manihot , Alelos , Cromossomos , Diploide , Haplótipos , Manihot/genética , Melhoramento Vegetal , Análise de Sequência de DNA , Transcriptoma
2.
Cells ; 11(11)2022 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-35681444

RESUMO

Membrane-enclosed organelle compartmentalization is not the only way by which cell processes are spatially organized. Phase separation is emerging as a new driver in the organization of membrane-less compartments and biological processes. Liquid-liquid phase separation has been indicated as a new way to control the kinetics of molecular reactions and is based on weak multivalent interactions affecting the stoichiometry of the molecules involved. In the nucleus, liquid-liquid phase separation may represent an ancestral means of controlling genomic activity by forming discrete chromatin regions, regulating transcriptional activity, contributing to the assembly of DNA damage response foci, and controlling the organization of chromosomes. Liquid-liquid phase separation also contributes to chromatin function through its role in the reorganization of the nuclear periphery in the post-mitotic phase. Herein, we describe the basic principles regulating liquid-liquid phase separation, analyze examples of phase separation occurring in the nucleus, and dedicate attention to the implication of liquid-liquid phase separation in the reorganization of the nuclear periphery by the endosomal sorting complexes required for transport (ESCRT) machinery. Although some caution is warranted, current scientific knowledge allows for the hypothesis that many factors and processes in the cell are yet to be discovered which are functionally associated with phase separation.


Assuntos
Núcleo Celular , Membrana Nuclear , Cromatina , Cromossomos , Complexos Endossomais de Distribuição Requeridos para Transporte/genética
4.
Sci Rep ; 12(1): 9699, 2022 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-35690623

RESUMO

Hodge theory reveals the deep intrinsic relations of differential forms and provides a bridge between differential geometry, algebraic topology, and functional analysis. Here we use Hodge Laplacian and Hodge decomposition models to analyze biomolecular structures. Different from traditional graph-based methods, biomolecular structures are represented as simplicial complexes, which can be viewed as a generalization of graph models to their higher-dimensional counterparts. Hodge Laplacian matrices at different dimensions can be generated from the simplicial complex. The spectral information of these matrices can be used to study intrinsic topological information of biomolecular structures. Essentially, the number (or multiplicity) of k-th dimensional zero eigenvalues is equivalent to the k-th Betti number, i.e., the number of k-th dimensional homology groups. The associated eigenvectors indicate the homological generators, i.e., circles or holes within the molecular-based simplicial complex. Furthermore, Hodge decomposition-based HodgeRank model is used to characterize the folding or compactness of the molecular structures, in particular, the topological associated domain (TAD) in high-throughput chromosome conformation capture (Hi-C) data. Mathematically, molecular structures are represented in simplicial complexes with certain edge flows. The HodgeRank-based average/total inconsistency (AI/TI) is used for the quantitative measurements of the folding or compactness of TADs. This is the first quantitative measurement for TAD regions, as far as we know.


Assuntos
Cromossomos , Análise de Dados , Estrutura Molecular
5.
Methods Mol Biol ; 2472: 95-108, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35674895

RESUMO

The sequence-specific transcription factor RBPJ, also known as CSL (CBF1, Su(H), Lag1), is an evolutionarily conserved protein that mediates Notch signaling to guide cell fates. When cells enter mitosis, DNA is condensed and most transcription factors dissociate from chromatin; however, a few, select transcription factors, termed bookmarking factors, remain associated. These mitotic chromatin-bound factors are believed to play important roles in maintaining cell fates through cell division. RBPJ is one such factor that remains mitotic chromatin associated and therefore could function as a bookmarking factor. Here, we describe how to obtain highly purified mitotic cells from the mouse embryonal carcinoma cell line F9, perform chromatin immunoprecipitation with mitotic cells, and measure the first run of RNA synthesis upon mitotic exit. These methods serve as basis to understand the roles of mitotic bookmarking by RBPJ in propagating Notch signals through cell division.


Assuntos
Cromatina , Cromossomos , Animais , Cromatina/genética , Cromossomos/metabolismo , Regulação da Expressão Gênica , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Camundongos , Mitose , Fatores de Transcrição/metabolismo
6.
Mol Biol Evol ; 39(6)2022 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-35642310

RESUMO

It is largely unknown how mammalian genomes evolve under rapid speciation and environmental adaptation. An excellent model for understanding fast evolution is provided by the genus Sus, which diverged relatively recently and lacks postzygotic isolation. Here, we present a high-quality reference genome of the Visayan warty pig, which is specialized to a tropical island environment. Comparing the genome sequences and chromatin contact maps of the Visayan warty pig (Sus cebifrons) and domestic pig (Sus scrofa), we characterized the dynamics of chromosomal structure evolution during Sus speciation, revealing the similar chromosome conformation as the potential biological mechanism of frequent postdivergence hybridization among Suidae. We further investigated the different signatures of adaptive selection and domestication in Visayan warty pig and domestic pig with specific emphasize on the evolution of olfactory and gustatory genes, elucidating higher olfactory diversity in Visayan warty pig and positive and relaxed evolution of bitter and fat taste receptors, respectively, in domestic pig. Our comprehensive evolutionary and comparative genome analyses provide insight into the dynamics of genomes and how these change over relative short evolutionary times, as well as how these genomic differences encode for differences in the phenotypes.


Assuntos
Cromossomos , Genoma , Animais , Genômica , Sus scrofa/genética , Suínos/genética
8.
Open Biol ; 12(6): 220062, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35642493

RESUMO

During cell division, drastic cellular changes characteristic of mitosis result in the inactivation of the transcriptional machinery, and global downregulation of transcription. Sequence-specific transcription factors (TFs) have thus been considered mere bystanders, devoid of any regulatory function during mitosis. This view changed significantly in recent years, upon the conclusion that many TFs associate with condensed chromosomes during cell division, even occupying a fraction of their genomic target sites in mitotic chromatin. This finding was at the origin of the concept of mitotic bookmarking by TFs, proposed as a mechanism to propagate gene regulatory information across cell divisions, by facilitating the reactivation of specific bookmarked genes. While the underlying mechanisms and biological significance of this model remain elusive, recent developments in this fast-moving field have cast new light into TF activity during mitosis, beyond a bookmarking role. Here, we start by reviewing the most recent findings on the complex nature of TF-chromatin interactions during mitosis, and on mechanisms that may regulate them. Next, and in light of recent reports describing how transcription is reinitiated in temporally distinct waves during mitosis-to-G1 transition, we explore how TFs may contribute to defining this hierarchical gene expression process. Finally, we discuss how TF activity during mitotic exit may impact the acquisition of cell identity upon cell division, and propose a model that integrates dynamic changes in TF-chromatin interactions during this cell-cycle period, with the execution of cell-fate decisions.


Assuntos
Mitose , Fatores de Transcrição , Cromatina/genética , Cromossomos/genética , Cromossomos/metabolismo , Regulação da Expressão Gênica , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
9.
Commun Biol ; 5(1): 608, 2022 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-35725901

RESUMO

Topologically associating domains (TADs) are fundamental building blocks of three dimensional genome, and organized into complex hierarchies. Identifying hierarchical TADs on Hi-C data helps to understand the relationship between genome architectures and gene regulation. Herein we propose TADfit, a multivariate linear regression model for profiling hierarchical chromatin domains, which tries to fit the interaction frequencies in Hi-C contact matrix with and without replicates using all-possible hierarchical TADs, and the significant ones can be determined by the regression coefficients obtained with the help of an online learning solver called Follow-The-Regularized-Leader (FTRL). Beyond the existing methods, TADfit has an ability to handle multiple contact matrix replicates and find partially overlapping TADs on them, which helps to find the comprehensive underlying TADs across replicates from different experiments. The comparative results tell that TADfit has better accuracy and reproducibility, and the hierarchical TADs called by it exhibit a reasonable biological relevance.


Assuntos
Cromatina , Cromossomos , Cromatina/genética , Genoma , Modelos Lineares , Reprodutibilidade dos Testes
10.
Planta ; 256(1): 12, 2022 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-35710953

RESUMO

MAIN CONCLUSIONS: Heat shock proteins, ROS detoxifying enzymes, and ion homeostasis proteins, together with proteins in carbohydrate metabolism, cell structure, brassinosteroids, and carotenoid biosynthesis pathway were up-regulated in CSSLs under salinity stress. Rice is one of the most consumed staple foods worldwide. Salinity stress is a serious global problem affecting rice productivity. Many attempts have been made to select or produce salinity-tolerant rice varieties. Genetics and biochemical approaches were used to study the salinity-responsive pathway in rice to develop salinity tolerant strains. This study investigated the proteomic profiles of chromosome segment substitution lines (CSSLs) developed from KDML105 (Khao Dawk Mali 105, a Thai jasmine rice cultivar) under salinity stress. The CSSLs showed a clear resistant phenotype in response to 150 mM NaCl treatment compared to the salinity-sensitive line, IR29. Liquid chromatography-tandem mass spectrometry using the Ultimate 3000 Nano/Capillary LC System coupled to a Hybrid Quadrupole Q-Tof Impact II™ equipped with a nano-captive spray ion source was applied for proteomic analysis. Based on our criteria, 178 proteins were identified as differentially expressed proteins under salinity stress. Protein functions in DNA replication and transcription, and stress and defense accounted for the highest proportions in response to salinity stress, followed by protein transport and trafficking, carbohydrate metabolic process, signal transduction, and cell structure. The protein interaction network among the 75 up-regulated proteins showed connections between proteins involved in cell wall synthesis, transcription, translation, and in defense responses.


Assuntos
Jasminum , Oryza , Cromossomos/metabolismo , Jasminum/genética , Jasminum/metabolismo , Oryza/genética , Oryza/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteômica , Salinidade , Estresse Salino/genética , Estresse Fisiológico/genética , Tailândia
11.
Gene ; 834: 146650, 2022 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-35680022

RESUMO

Taenia pisiformis is one of the most widespread gastrointestinal parasites and its larvae (cysticercosis) causes significant economic loss to rabbit industry. No efficient drug is available for this disease to date. To better understand its genomics, we assembled a 211-Mb high quality genome of T. pisiformis at chromosome level with a scaffold N50 size of 20 Mbp. Totally, 12,097 protein-coding genes was predicted from the genome. Genome-level phylogenetic analysis confirmed the taxonomic affiliations with other tapeworms and revealed that T. pisiformis diverged from its closely related relative T. hydatigena âˆ¼ 14.6 Mya. Comparative genomic analyses revealed that the T. pisiformis genome was characterized by adaptive features of strong positive selection signals from carbohydrate/lipid metabolism and body surface integrity, and of expanded gene families related to metabolism of amino acids and lipids. The high-quality genome of T. pisiformis constitutes a resource for the comparative genomics and for further applications in general parasitology.


Assuntos
Cestoides , Taenia , Animais , Cestoides/genética , Cromossomos/genética , Filogenia , Coelhos , Taenia/genética
12.
Mol Cell ; 82(12): 2350-2350.e1, 2022 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-35714589

RESUMO

Chromosomes in higher eukaryotes are folded at different length scales into loop extrusion domains, spatial compartments, and chromosome territories and exhibit interactions with nuclear structures such as the lamina. Microscopic methods can probe this structure by measuring positions of chromosomes in the nuclear space in individual cells, while sequencing-based contact capture approaches can report the frequency of contacts of different regions within these structural layers. To view this SnapShot, open or download the PDF.


Assuntos
Cromatina , Cromossomos , Núcleo Celular/genética , Núcleo Celular/ultraestrutura , Cromossomos/genética , Eucariotos/genética
13.
Nat Commun ; 13(1): 3488, 2022 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-35715427

RESUMO

The expression of some genes depends on large, adjacent regions of the genome that contain multiple enhancers. These regulatory landscapes frequently align with Topologically Associating Domains (TADs), where they integrate the function of multiple similar enhancers to produce a global, TAD-specific regulation. We asked if an individual enhancer could overcome the influence of one of these landscapes, to drive gene transcription. To test this, we transferred an enhancer from its native location, into a nearby TAD with a related yet different functional specificity. We used the biphasic regulation of Hoxd genes during limb development as a paradigm. These genes are first activated in proximal limb cells by enhancers located in one TAD, which is then silenced when the neighboring TAD activates its enhancers in distal limb cells. We transferred a distal limb enhancer into the proximal limb TAD and found that its new context suppresses its normal distal specificity, even though it is bound by HOX13 transcription factors, which are responsible for the distal activity. This activity can be rescued only when a large portion of the surrounding environment is removed. These results indicate that, at least in some cases, the functioning of enhancer elements is subordinated to the host chromatin context, which can exert a dominant control over its activity.


Assuntos
Cromatina , Elementos Facilitadores Genéticos , Cromatina/genética , Cromossomos , Elementos Facilitadores Genéticos/genética , Extremidades , Fatores de Transcrição/genética
14.
Nat Commun ; 13(1): 3511, 2022 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-35717499

RESUMO

Patchouli (Pogostemon cablin (Blanco) Benth.), a member of the Lamiaceae family, is an important aromatic plant that has been widely used in medicine and perfumery. Here, we report a 1.94 Gb chromosome-scale assembly of the patchouli genome (contig N50 = 7.97 Mb). The gene annotation reveals that tandem duplication of sesquiterpene biosynthetic genes may be a major contributor to the biosynthesis of patchouli bioactivity components. We further phase the genome into two distinct subgenomes (A and B), and identify a chromosome substitution event that have occurred between them. Further investigations show that a burst of universal LTR-RTs in the A subgenome lead to the divergence between two subgenomes. However, no significant subgenome dominance is detected. Finally, we track the evolutionary scenario of patchouli including whole genome tetraploidization, subgenome divergency, hybridization, and chromosome substitution, which are the key forces to determine the complexity of patchouli genome. Our work sheds light on the evolutionary history of patchouli and offers unprecedented genomic resources for fundamental patchouli research and elite germplasm development.


Assuntos
Lamiaceae , Pogostemon , Cromossomos , Haplótipos , Lamiaceae/genética , Pogostemon/genética , Tetraploidia
15.
Zoolog Sci ; 39(3): 253-260, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35699928

RESUMO

Gene/transcript model sets predicted from decoded genome sequences are an important resource for a wide range of biological studies. Accuracy of gene models is therefore critical for deducing accurate conclusions. Computationally predicted models are sometimes inconsistent with experimental data from cDNA clones and RNA-sequencing. In an ascidian, Ciona robusta (Ciona intestinalis type A), a manually curated gene/transcript model set, which was constructed using an assembly in which 68% of decoded sequences were associated with chromosomes, had been used during the last decade. Recently a new genome assembly was published, in which over 95% of decoded sequences are associated with chromosomes. In the present study, we provide a high-quality version of the gene/transcript model set for the latest assembly. Because the Ciona genome has been used in a variety of studies such as developmental biological studies, evolutionary studies, and physiological studies, the current gene/transcript model set provides a fundamental biological resource.


Assuntos
Ciona intestinalis , Animais , Sequência de Bases , Evolução Biológica , Cromossomos , Ciona intestinalis/genética , Genoma
16.
Gigascience ; 112022 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-35701375

RESUMO

BACKGROUND: Bretschneidera sinensis is an endangered relic tree species in the Akaniaceae family and is sporadically distributed in eastern Asia. As opposed to its current narrow and rare distribution, the fossil pollen of B. sinensis has been found to be frequent and widespread in the Northern Hemisphere during the Late Miocene. B. sinensis is also a typical mycorrhizal plant, and its annual seedlings exhibit high mortality rates in absence of mycorrhizal development. The chromosome-level high-quality genome of B. sinensis will help us to more deeply understand the survival and demographic histories of this relic species. RESULTS: A total of 25.39 Gb HiFi reads and 109.17 Gb Hi-C reads were used to construct the chromosome-level genome of B. sinensis, which is 1.21 Gb in length with the contig N50 of 64.13 Mb and chromosome N50 of 146.54 Mb. The identified transposable elements account for 55.21% of the genome. A total of 45,839 protein-coding genes were predicted in B. sinensis. A lineage-specific whole-genome duplication was detected, and 7,283 lineage-specific expanded gene families with functions related to the specialized endotrophic mycorrhizal adaptation were identified. The historical effective population size (Ne) of B. sinensis was found to oscillate greatly in response to Quaternary climatic changes. The Ne of B. sinensis has decreased rapidly in the recent past, making its extant Ne extremely lower. Our additional evolutionary genomic analyses suggested that the developed mycorrhizal adaption might have been repeatedly disrupted by environmental changes caused by Quaternary climatic oscillations. The environmental changes and an already decreased population size during the Holocene may have led to the current rarity of B. sinensis. CONCLUSION: This is a detailed report of the genome sequences for the family Akaniaceae distributed in evergreen forests in eastern Asia. Such a high-quality genomic resource may provide critical clues for comparative genomics studies of this family in the future.


Assuntos
Genoma , Magnoliopsida , Animais , Cromossomos , Demografia , Espécies em Perigo de Extinção , Genômica , Filogenia
17.
Gigascience ; 112022 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-35701376

RESUMO

Artocarpus nanchuanensis (Moraceae), which is naturally distributed in China, is a representative and extremely endangered tree species. In this study, we obtained a high-quality chromosome-scale genome assembly and annotation information for A. nanchuanensis using integrated approaches, including Illumina, Nanopore sequencing platform, and Hi-C. A total of 128.71 Gb of raw Nanopore reads were generated from 20-kb libraries, and 123.38 Gb of clean reads were obtained after filtration with 160.34× coverage depth and a 17.48-kb average read length. The final assembled A. nanchuanensis genome was 769.44 Mb with a 2.09 Mb contig N50, and 99.62% (766.50 Mb) of the assembled data was assigned to 28 pseudochromosomes. In total, 39,596 genes (95.10%, 39,596/41,636) were successfully annotated, and 129 metabolic pathways were detected. Plants disease resistance/insect resistance genes, plant-pathogen interaction metabolic pathways, and abundant biosynthesis pathways of vitamins, flavonoid, and gingerol were detected. Unigene reveals the basis of species-specific functions, and gene family in contraction and expansion generally implies strong functional differences in the evolution. Compared with other related species, a total of 512 unigenes, 309 gene families in contraction, and 559 gene families in expansion were detected in A. nanchuanensis. This A. nanchuanensis genome information provides an important resource to expand our understanding of the unique biological processes, nutritional and medicinal benefits, and evolutionary relationship of this species. The study of gene function and metabolic pathway in A. nanchuanensis may reveal the theoretical basis of a special trait in A. nanchuanensis and promote the study and utilization of its rare medicinal value.


Assuntos
Artocarpus , Moraceae , Artocarpus/genética , Cromossomos , Frutas , Anotação de Sequência Molecular , Moraceae/genética , Filogenia , Árvores/genética
18.
BMC Biol ; 20(1): 139, 2022 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-35698132

RESUMO

BACKGROUND: Peach (Prunus persica) is an economically important stone fruit crop in Rosaceae and widely cultivated in temperate and subtropical regions, emerging as an excellent material to study the interaction between plant and environment. During its genus, there are four wild species of peach, all living in harsh environments. For example, one of the wild species, P. mira, originates from the Qinghai-Tibet Plateau (QTP) and exhibits strong cold/ultraviolet ray environmental adaptations. Although remarkable progresses in the gene discovery of fruit quality-related traits in peach using previous assembled genome were obtained, genomic basis of the response of these wild species to different geographical environments remains unclear. RESULTS: To uncover key genes regulating adaptability in different species and analyze the role of genetic variations in resistance formation, we performed de novo genome assembling of four wild relatives of peach (P. persica), P. mira, P. davidiana, P. kansuensis, and P. ferganensis and resequenced 175 peach varieties. The phylogenetic tree showed that the divergence time of P. mira and other wild relatives of peach was 11.5 million years ago, which was consistent with the drastic crustal movement of QTP. Abundant genetic variations were identified in four wild species when compared to P. persica, and the results showed that plant-pathogen interaction pathways were enriched in genes containing small insertions and deletions and copy number variations in all four wild relatives of peach. Then, the data were used to identify new genes and variations regulating resistance. For example, presence/absence variations which result from a hybridization event that occurred between P. mira and P. dulcis enhanced the resistance of their putative hybrid, P. davidiana. Using bulked segregant analysis, we located the nematode resistance locus of P. kansuensis in chromosome 2. Within the mapping region, a deletion in the promoter of one NBS-LRR gene was found to involve the resistance by regulating gene expression. Furthermore, combined with RNA-seq and selective sweeps analysis, we proposed that a deletion in the promoter of one CBF gene was essential for high-altitude adaptation of P. mira through increasing its resistance to low temperature. CONCLUSIONS: In general, the reference genomes assembled in the study facilitate our understanding of resistance mechanism of perennial fruit crops, and provide valuable resources for future breeding and improvement.


Assuntos
Prunus persica , Cromossomos , Variações do Número de Cópias de DNA , Evolução Molecular , Genoma de Planta , Filogenia , Melhoramento Vegetal , Prunus persica/genética
19.
Biosystems ; 218: 104706, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-35643186

RESUMO

The existence of an environmentally regulated version of meiotic crossing-over, or eco-crossover, is proposed, and the main consequences of this hypothesis are considered. Eco-crossover is a key source of partially directed genetic diversity of eukaryotes. In stressful environment, it creates ecologically justified and topologically specific genetic changes, and hence phenotypic variability, with which the selection works. If variability were random, then, in the face of rapid environmental changes, natural selection could not create life-saving adaptations in a timely manner. Owing to the eco-crossover activity, epimutations, i.e., eco-dependently marked chromosomal sites, are transforming into mutations. In its work, eco-crossover uses the eco-stress-dependent versions of circular RNAs ("ecological" circRNAs), which, against the background of eco-stresses, are synthesized as variants of alternative splicing. These ecological circRNAs, binding to homologous epimutations on the homologous parent chromosomes of the meiocyte, involve them in topologically specific recombinations. These recombinations can create random mutations in nonrandom genomic sites. These quasi-random mutations serve as a pivotal source for creating all adaptations of any level of complexity. The drivers of the adaptive evolution of eukaryotes, both in micro- and macroevolution, are two irreplaceable factors - eco-crossover and natural selection.


Assuntos
Troca Genética , RNA Circular , Cromossomos , Troca Genética/genética , Eucariotos/genética , Genoma , Meiose , Seleção Genética
20.
J Vis Exp ; (183)2022 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-35660707

RESUMO

Principles of DNA folding in the cell nucleus and its dynamic transformations that occur during the fulfillment of basic genetic functions (transcription, replication, segregation, etc.) remain poorly understood, partially due to the lack of experimental approaches to high-resolution visualization of specific chromatin loci in structurally preserved nuclei. Here we present a protocol for the visualization of replicative domains in monolayer cell culture in situ, by combining EdU labeling of newly synthesized DNA with subsequent label detection with Ag-amplification of Nanogold particles and ChromEM staining of chromatin. This protocol allows for the high-contrast, high-efficiency pre-embedding labeling, compatible with traditional glutaraldehyde fixation that provides the best structural preservation of chromatin for room-temperature sample processing. Another advantage of pre-embedding labeling is the possibility to pre-select cells of interest for sectioning. This is especially important for the analysis of heterogeneous cell populations, as well as compatibility with electron tomography approaches to high-resolution 3D analysis of chromatin organization at sites of replication, and the analysis of post-replicative chromatin rearrangement and sister chromatid segregation in the interphase.


Assuntos
Cromatina , Tomografia com Microscopia Eletrônica , Núcleo Celular/genética , Cromatina/genética , Cromossomos , DNA/química , Interfase
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