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Fatty acid esters obtained as a result of transesterification of vegetable oils or chemical reactions with alcohols are currently the main alternative fuels used in power plants. Since they do not contain aromatic hydrocarbons and sulfur compounds, emissions of CO2, hydrocarbons, carcinogens and carcinogenic substances into the atmosphere are significantly reduced compared to fuels derived from oil. An analysis of the efficiency reserves of the most promising biodiesel production technologies shows the advantages of using supercritical fluids to isolate CO2 from a biodiesel mixture and then separate glycerol, which is associated with environmental cleanliness, safety and energy efficiency. It is shown that the improvement of the technology of supercritical fluids for the production of biodiesel from vegetable oil can be achieved by connecting the OCF absorption-water-ammonia refrigeration unit technology to the general energy supply scheme.

Algebraic quasi-fractal is a convenient form of recording a process. Currently, digital transformation’s process permeates almost all areas of human activity. In this regard, digital twins of risk management methods become necessary. In this article, we approach the study of the risk system of a complex system from the following position: a risk event in the system occurs in the event of an intermission of the system's connection. The main idea to describe the risk system of a complex system is the description as model with intermission connections. The main tool for it the algebraic quasi – fractal systems. So, it will be used: modeling of the risk system of a complex system in accordance with a model in the form of a group of factors determining the complex system, with a help of a group with an intermission a definite connection of a system; modeling of the risk system of a complex system in accordance with a quasi – fractal group of factors determining the complex system, with a help of a quasi – fractal group with an intermission a definite quasi – fractal connection of a complex system. Such approach allows one to connect properties of a complex system with properties of its risk system.

Heat pump technology for utilisation of heat of secondary energy resources in catalytic method of production of biodiesel fuel is implemented. Algorithm for control of technological parameters at all stages of production of biodiesel fuel is proposed and analysis of parameters of its quality is performed. Heat pump system is represented in the form of two recirculation circuits of low-potential and high-potential heat carriers, preparation of which was carried out in stages of low and high pressure of heat pump. Exergetic analysis was performed, functional dependencies of exergy flows coming out of the considered types of process equipment were obtained. The obtained hybesel fuel complied with the requirements of GOST 23513-79. The proposed heat pump technology reduced specific energy consumption by 13-14%; improve environmental safety at all stages of the process; maximum reduction of spent heat carriers emission into the ambient atmosphere.

The tourist destinations develop with the participation of a significant number of stakeholders, whose interests often do not coincide, and sometimes completely differently directed, which determines the multifactorial nature of the relationship in the scope of the set goals achieving. On the one hand, this is the development of tourism, in which the business community is interested, on the other hand, the interests of the territory administration, which interests are of a dual nature. Coordination of the interests of all interested parties - the administration, the population of the territorial entity, the business community and tourists becomes a priority in this situation. The solution to this priority task through partnership management technologies is proposed in this research. The research is based on the use of a systematic approach and technologies of management analysis and modelling. The proposed conclusion, algorithm, model can be used in the implementation of territories branding, in the formation and implementation of the tourist destination development, in the regional policy formulation in the tourism development and the local self-government development.

The subgenus Stenocranius contains two cryptic species: Lasiopodomys gregalis (subdivided into three allopatrically distributed and genetically well-isolated lineages A, B, and C) and Lasiopodomys raddei. To identify karyotype characteristics of this poorly studied cryptic species complex, we used comparative cytogenetic analysis of 138 individuals from 41 localities in South Siberia and Mongolia. A detailed description of the L. raddei karyotype and of the L. gregalis lineage С karyotype is presented for the first time. The A chromosome complement of all examined narrow-headed voles consisted of 2n = 36 and a fundamental number of autosomal arms (FNa) of 50. Between species, patterns of differential staining were similar, though additional C-heterochromatic blocks were found in L. gregalis lineages; Ag-positive nucleolar organizers and ribosomal DNA (rDNA) clusters are located on eight and nine acrocentric pairs, respectively. No B chromosomes (Bs) were found in the Early Pleistocene relic L. raddei, while one to five small heterochromatic acrocentric Bs were detected in all L. gregalis lineages; the number and frequency of Bs varied considerably within lineages, but no intraindividual variation was observed. In both species, telomeric repeats were visualized at termini of all chromosomes, including Bs. The number and localization of rDNA clusters on Bs varied among B-carriers. Immunodetection of several meiotic proteins indicated that meio-Bs are transcriptionally inactive and have a pattern of meiotic behavior similar to that of sex chromosomes (some homology of Bs to sex chromosomes is supposed). The nature, mechanisms of inheritance and stability of Bs in L. gregalis require further investigation., (© 2024 Wiley Periodicals LLC.)

Analyses of ancient DNA typically involve sequencing the surviving short oligonucleotides and aligning to genome assemblies from related, modern species. Here, we report that skin from a female woolly mammoth (†Mammuthus primigenius) that died 52,000 years ago retained its ancient genome architecture. We use PaleoHi-C to map chromatin contacts and assemble its genome, yielding 28 chromosome-length scaffolds. Chromosome territories, compartments, loops, Barr bodies, and inactive X chromosome (Xi) superdomains persist. The active and inactive genome compartments in mammoth skin more closely resemble Asian elephant skin than other elephant tissues. Our analyses uncover new biology. Differences in compartmentalization reveal genes whose transcription was potentially altered in mammoths vs. elephants. Mammoth Xi has a tetradic architecture, not bipartite like human and mouse. We hypothesize that, shortly after this mammoth's death, the sample spontaneously freeze-dried in the Siberian cold, leading to a glass transition that preserved subfossils of ancient chromosomes at nanometer scale., Competing Interests: Declaration of interests E.L.A., M.T.P.G., and L.D. are on the scientific advisory board of Colossal Biosciences and hold stock options. From 2021 to 2023, M.A.M.-R. received consulting honoraria from Acuity Spatial Genomics. E.L.A. and O.D. are inventors on US provisional patent applications 16/308,386 (E.L.A. and O.D., filed 12/7/2018), 16/247,502 (E.L.A. and O.D., 1/14/2019), and PCT/US2020/064704 (E.L.A., 12/11/2020) by the Baylor College of Medicine and the Broad Institute, relating to the assembly methods in this manuscript., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Descriptions of karyotypes of many animal species are currently available. In addition, there has been a significant increase in the number of sequenced genomes and an ever-improving quality of genome assembly. To close the gap between genomic and cytogenetic data we applied fluorescent in situ hybridization (FISH) and Hi-C technology to make the first full chromosome-level genome comparison of the guinea pig (Cavia porcellus), naked mole-rat (Heterocephalus glaber), and human. Comparative chromosome maps obtained by FISH with chromosome-specific probes link genomic scaffolds to individual chromosomes and orient them relative to centromeres and heterochromatic blocks. Hi-C assembly made it possible to close all gaps on the comparative maps and to reveal additional rearrangements that distinguish the karyotypes of the three species. As a result, we integrated the bioinformatic and cytogenetic data and adjusted the previous comparative maps and genome assemblies of the guinea pig, naked mole-rat, and human. Syntenic associations in the two hystricomorphs indicate features of their putative ancestral karyotype. We postulate that the two approaches applied in this study complement one another and provide complete information about the organization of these genomes at the chromosome level., (© 2023. The Author(s).)

Electric fields are now considered a major mechanism of epileptiform activity. However, it is not clear if another electrophysiological phenomenon, burst suppression, utilizes the same mechanism for its bursting phase. Thus, the purpose of this study was to compare the role of ephaptic coupling-the recruitment of neighboring cells via electric fields-in generating bursts in epilepsy and burst suppression. We used local injections of the GABA-antagonist picrotoxin to elicit epileptic activity and a general anesthetic, sevoflurane, to elicit burst suppression in rabbits. Then, we applied an established computational model of pyramidal cells to simulate neuronal activity in a 3-dimensional grid, with an additional parameter to trigger a suppression phase based on extra-cellular calcium dynamics. We discovered that coupling via electric fields was sufficient to produce bursting in scenarios where inhibitory control of excitatory neurons was sufficiently low. Under anesthesia conditions, bursting occurs with lower neuronal recruitment in comparison to seizures. Our model predicts that due to the effect of electric fields, the magnitude of bursts during seizures should be roughly 2-3 times the magnitude of bursts that occur during burst suppression, which is consistent with our in vivo experimental results. The resulting difference in magnitude between bursts during anesthesia and epileptiform bursts reflects the strength of the electric field effect, which suggests that burst suppression and epilepsy share the same ephaptic coupling mechanism.

The functional deficiency of the inhibitory system typically appears during development and can progress to psychiatric disorders or epilepsy, depending on its severity, in later years. It is known that interneurons, the major source of GABAergic inhibition in the cerebral cortex, can make direct connections with arterioles and participate in the regulation of vasomotion. The goal of this study was to mimic the functional deficiency of interneurons through the use of localized microinjections of the GABA antagonist, picrotoxin, in such a concentration that it did not elicit epileptiform neuronal activity. First, we recorded the dynamics of resting-state neuronal activity in response to picrotoxin injections in the somatosensory cortex of an awake rabbit; second, we assessed the altered neuronal and hemodynamic responses to whisker stimulation using BOLD fMRI and electrophysiology recordings; third, we evaluated brain tissue oxygen levels before and after picrotoxin injection. Our results showed that neuronal activity typically increased after picrotoxin administration, the BOLD responses to stimulation became negative, and the oxygen response was nearly abolished. Vasoconstriction during the resting baseline was not observed. These results indicate that picrotoxin provoked imbalanced hemodynamics either due to increased neuronal activity, decreased vascular response, or a combination of both.

Pusa sibirica , the Baikal seal, is the only extant, exclusively freshwater, pinniped species. The pending issue is, how and when they reached their current habitat-the rift lake Baikal, more than three thousand kilometers away from the Arctic Ocean. To explore the demographic history and genetic diversity of this species, we generated a de novo chromosome-length assembly, and compared it with three closely related marine pinniped species. Multiple whole genome alignment of the four species compared with their karyotypes showed high conservation of chromosomal features, except for three large inversions on chromosome VI. We found the mean heterozygosity of the studied Baikal seal individuals was relatively low (0.61 SNPs/kbp), but comparable to other analyzed pinniped samples. Demographic reconstruction of seals revealed differing trajectories, yet remarkable variations in Ne occurred during approximately the same time periods. The Baikal seal showed a significantly more severe decline relative to other species. This could be due to the difference in environmental conditions encountered by the earlier populations of Baikal seals, as ice sheets changed during glacial-interglacial cycles. We connect this period to the time of migration to Lake Baikal, which occurred ~3-0.3 Mya, after which the population stabilized, indicating balanced habitat conditions.

Constitutive-heterochromatin placement in the genome affects chromosome structure by occupying centromeric areas and forming large blocks. To investigate the basis for heterochromatin variation in the genome, we chose a group of species with a conserved euchromatin part: the genus Martes [stone marten ( M. foina, 2n = 38), sable ( M. zibellina , 2n = 38 ) , pine marten ( M. martes, 2n = 38), and yellow-throated marten ( M. flavigula , 2n = 40)]. We mined the stone marten genome for the most abundant tandem repeats and selected the top 11 macrosatellite repetitive sequences. Fluorescent in situ hybridization revealed distributions of the tandemly repeated sequences (macrosatellites, telomeric repeats, and ribosomal DNA). We next characterized the AT/GC content of constitutive heterochromatin by CDAG (Chromomycin A3-DAPI-after G-banding). The euchromatin conservatism was shown by comparative chromosome painting with stone marten probes in newly built maps of the sable and pine marten. Thus, for the four Martes species, we mapped three different types of tandemly repeated sequences critical for chromosome structure. Most macrosatellites are shared by the four species with individual patterns of amplification. Some macrosatellites are specific to a species, autosomes, or the X chromosome. The variation of core macrosatellites and their prevalence in a genome are responsible for the species-specific variation of the heterochromatic blocks.

Localized increases in neuronal activity are supported by the hemodynamic response, which delivers oxygen to the brain tissue to support synaptic functions, action potentials and other neuronal processes. However, it remains unknown if changes in baseline neuronal activity, which are expected to reflect neuronal metabolic demand, alter the relationship between the local hemodynamic and oxygen behaviour. In order to better characterize this system, we examine here the relationship between brain tissue oxygen (PO 2 ) and hemodynamic responses (BOLD functional MRI) under different levels of neuronal activity. By comparing the stimulus-evoked responses during different levels of baseline neuronal activity, the awake state vs isoflurane anesthesia, we were able to measure how a known change in neuronal demand affected tissue PO 2 as well as the hemodynamic response to stimulation. We observed a high correlation between stimulus-evoked PO 2 and BOLD responses in the awake state. Moreover, we found that the evoked PO 2 and BOLD responses were still present despite the elevated tissue oxygen baseline and decreased baseline of neuronal activity under low concentration isoflurane, and that the magnitudes of these responses decreased by similar proportions but the relationship between these signals was distorted. Our findings point to distortion of the BOLD-PO 2 relationship due to anesthesia. The feedback mechanism to adjust the level of brain tissue oxygen, as well as the correlation between BOLD and PO 2 responses, are impaired even by a small dose of anesthetics., (© 2023. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

The dynamic interaction between excitatory and inhibitory activity in the brain is known as excitatory-inhibitory balance (EIB). A significant shift in EIB toward excitation has been observed in numerous pathological states and diseases, such as autism or epilepsy, where interneurons may be dysfunctional. The consequences of this on neurovascular interactions remains to be elucidated. Specifically, it is not known if there is an elevated metabolic consumption of oxygen due to increased excitatory activity. To investigate this, we administered microinjections of picrotoxin, a gamma aminobutyric acid (GABA) antagonist, to the rabbit cortex in the awake state to mimic the functional deficiency of GABAergic interneurons. This caused an observable shift in EIB toward excitation without the induction of seizures. We used chronically implanted electrodes to measure both neuronal activity and brain tissue oxygen concentrations (PO 2 ) simultaneously and in the same location. Using a high-frequency recording rate for PO 2 , we were able to detect two important phenomena, (1) the shift in EIB led to a change in the power spectra of PO 2 fluctuations, such that higher frequencies (8-15 cycles per minute) were suppressed and (2) there were brief periods (dips with a duration of less than 100 ms associated with neuronal bursts) when PO 2 dropped below 10 mmHg, which we defined as the threshold for hypoxia. The dips were followed by an overshoot, which indicates either a rapid vascular response or decrease in oxygen consumption. Our results point to the essential role of interneurons in brain tissue oxygen regulation in the resting state., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Aksenov, Doubovikov, Serdyukova, Gascoigne, Linsenmeier and Drobyshevsky.)

Human and animal studies have elucidated the apparent neurodevelopmental effects resulting from neonatal anesthesia. Observations of learning and behavioral deficits in children, who were exposed to anesthesia early in development, have instigated a flurry of studies that have predominantly utilized animal models to further interrogate the mechanisms of neonatal anesthesia-induced neurotoxicity. Specifically, while neonatal anesthesia has demonstrated its propensity to affect multiple cell types in the brain, it has shown to have a particularly detrimental effect on the gamma aminobutyric acid (GABA)ergic system, which contributes to the observed learning and behavioral deficits. The damage to GABAergic neurons, resulting from neonatal anesthesia, seems to involve structure-specific changes in excitatory-inhibitory balance and neurovascular coupling, which manifest following a significant interval after neonatal anesthesia exposure. Thus, to better understand how neonatal anesthesia affects the GABAergic system, we first review the early development of the GABAergic system in various structures that have been the focus of neonatal anesthesia research. This is followed by an explanation that, due to the prolonged developmental curve of the GABAergic system, the entirety of the negative effects of neonatal anesthesia on learning and behavior in children are not immediately evident, but instead take a substantial amount of time (years) to fully develop. In order to address these concerns going forward, we subsequently offer a variety of in vivo methods which can be used to record these delayed effects.

The Puma lineage within the family Felidae consists of 3 species that last shared a common ancestor around 4.9 million years ago. Whole-genome sequences of 2 species from the lineage were previously reported: the cheetah (Acinonyx jubatus) and the mountain lion (Puma concolor). The present report describes a whole-genome assembly of the remaining species, the jaguarundi (Puma yagouaroundi). We sequenced the genome of a male jaguarundi with 10X Genomics linked reads and assembled the whole-genome sequence. The assembled genome contains a series of scaffolds that reach the length of chromosome arms and is similar in scaffold contiguity to the genome assemblies of cheetah and puma, with a contig N50 = 100.2 kbp and a scaffold N50 = 49.27 Mbp. We assessed the assembled sequence of the jaguarundi genome using BUSCO, aligned reads of the sequenced individual and another published female jaguarundi to the assembled genome, annotated protein-coding genes, repeats, genomic variants and their effects with respect to the protein-coding genes, and analyzed differences of the 2 jaguarundis from the reference mitochondrial genome. The jaguarundi genome assembly and its annotation were compared in quality, variants, and features to the previously reported genome assemblies of puma and cheetah. Computational analyzes used in the study were implemented in transparent and reproducible way to allow their further reuse and modification., (© The American Genetic Association. 2021.)

The taxonomy of the genus Calomyscus remains controversial. According to the latest systematics the genus includes eight species with great karyotypic variation. Here, we studied karyotypes of 14 Calomyscus individuals from different regions of Iran and Turkmenistan using a new set of chromosome painting probes from a Calomyscus sp. male (2n = 46, XY; Shahr-e-Kord-Soreshjan-Cheshme Maiak Province). We showed the retention of large syntenic blocks in karyotypes of individuals with identical chromosome numbers. The only rearrangement (fusion 2/21) differentiated Calomyscus elburzensis, Calomyscus mystax mystax , and Calomyscus sp. from Isfahan Province with 2n = 44 from karyotypes of C. bailwardi, Calomyscus sp. from Shahr-e-Kord, Chahar Mahal and Bakhtiari-Aloni, and Khuzestan-Izeh Provinces with 2n = 46. The individuals from Shahdad tunnel, Kerman Province with 2n = 51-52 demonstrated non-centric fissions of chromosomes 4, 5, and 6 of the 46-chromosomal form with the formation of separate small acrocentrics. A heteromorphic pair of chromosomes in a specimen with 2n = 51 resulted from a fusion of two autosomes. C-banding and chromomycin A3-DAPI staining after G-banding showed extensive heterochromatin variation between individuals.

The genus status of Urocricetus was defined recently based on morphological and molecular data. Even though the amount of evidence for a separate phylogenetic position of this genus among Cricetinae continues to increase, there is still no consensus on its relationship to other groups. Here we give the first comprehensive description of the U. kamensis karyotype (2n = 30, NFa = 50) including results of comparative cytogenetic analysis and detailed examination of its phylogenetic position by means of numerous molecular markers. The molecular data strongly indicated that Urocricetus is a distant sister group to Phodopus. Comparative cytogenetic data showed significant reorganization of the U. kamensis karyotype compared to karyotypes of all other hamsters investigated earlier. The totality of findings undoubtedly means that Urocricetus belongs to a separate divergent lineage of Cricetinae.

Pinnipedia karyotype evolution was studied here using human, domestic dog, and stone marten whole-chromosome painting probes to obtain comparative chromosome maps among species of Odobenidae ( Odobenus rosmarus ), Phocidae ( Phoca vitulina , Phoca largha , Phoca hispida , Pusa sibirica , Erignathus barbatus ), and Otariidae ( Eumetopias jubatus , Callorhinus ursinus , Phocarctos hookeri , and Arctocephalus forsteri ). Structural and functional chromosomal features were assessed with telomere repeat and ribosomal-DNA probes and by CBG (C-bands revealed by barium hydroxide treatment followed by Giemsa staining) and CDAG (Chromomycin A3-DAPI after G-banding) methods. We demonstrated diversity of heterochromatin among pinniped karyotypes in terms of localization, size, and nucleotide composition. For the first time, an intrachromosomal rearrangement common for Otariidae and Odobenidae was revealed. We postulate that the order of evolutionarily conserved segments in the analyzed pinnipeds is the same as the order proposed for the ancestral Carnivora karyotype (2n = 38). The evolution of conserved genomes of pinnipeds has been accompanied by few fusion events (less than one rearrangement per 10 million years) and by novel intrachromosomal changes including the emergence of new centromeres and pericentric inversion/centromere repositioning. The observed interspecific diversity of pinniped karyotypes driven by constitutive heterochromatin variation likely has played an important role in karyotype evolution of pinnipeds, thereby contributing to the differences of pinnipeds' chromosome sets.

Euchromatic segments of the X chromosomes of placental mammals are the most conservative elements of the karyotype, only rarely subjected to either inter- or intrachromosomal rearrangements. Here, using microdissection-derived set of region-specific probes of Terricola savii we detailed the evolutionary rearrangements found in X chromosomes in 20 vole species (Arvicolinae, Rodentia). We show that the evolution of X chromosomes in this taxon was accompanied by multiple para- and pericentric inversions and centromere shifts. The contribution of intrachromosomal rearrangements to the karyotype evolution of Arvicolinae species was approximately equivalent in both the separate autosomal conserved segments and the X chromosomes. Intrachromosmal rearrangements and structural reorganization of the X chromosomes was likely accompanied by an accumulation, distribution, and evolution of repeated sequences.

The history of each human chromosome can be studied through comparative cytogenetic approaches in mammals which permit the identification of human chromosomal homologies and rearrangements between species. Comparative banding, chromosome painting, Bacterial Artificial Chromosome (BAC) mapping and genome data permit researchers to formulate hypotheses about ancestral chromosome forms. Human chromosome 13 has been previously shown to be conserved as a single syntenic element in the Ancestral Primate Karyotype; in this context, in order to study and verify the conservation of primate chromosomes homologous to human chromosome 13, we mapped a selected set of BAC probes in three platyrrhine species, characterised by a high level of rearrangements, using fluorescence in situ hybridisation (FISH). Our mapping data on Saguinus oedipus, Callithrix argentata and Alouatta belzebul provide insight into synteny of human chromosome 13 evolution in a comparative perspective among primate species, showing rearrangements across taxa. Furthermore, in a wider perspective, we have revised previous cytogenomic literature data on chromosome 13 evolution in eutherian mammals, showing a complex origin of the eutherian mammal ancestral karyotype which has still not been completely clarified. Moreover, we analysed biomedical aspects (the OMIM and Mitelman databases) regarding human chromosome 13, showing that this autosome is characterised by a certain level of plasticity that has been implicated in many human cancers and diseases.

The mandarin vole, Lasiopodomys mandarinus , is one of the most intriguing species among mammals with non-XX/XY sex chromosome system. It combines polymorphism in diploid chromosome numbers, variation in the morphology of autosomes, heteromorphism of X chromosomes, and several sex chromosome systems the origin of which remains unexplained. Here we elucidate the sex determination system in Lasiopodomys mandarinus vinogradovi using extensive karyotyping, crossbreeding experiments, molecular cytogenetic methods, and single chromosome DNA sequencing. Among 205 karyotyped voles, one male and three female combinations of sex chromosomes were revealed. The chromosome segregation pattern and karyomorph-related reproductive performances suggested an aberrant sex determination with almost half of the females carrying neo-X/neo-Y combination. The comparative chromosome painting strongly supported this proposition and revealed the mandarin vole sex chromosome systems originated due to at least two de novo autosomal translocations onto the ancestral X chromosome. The polymorphism in autosome 2 was not related to sex chromosome variability and was proved to result from pericentric inversions. Sequencing of microdissection derived of sex chromosomes allowed the determination of the coordinates for syntenic regions but did not reveal any Y-specific sequences. Several possible sex determination mechanisms as well as interpopulation karyological differences are discussed.

The involvement of chromosome changes in the initial steps of speciation is controversial. Here we examine diversification trends within the mole voles Ellobius , a group of subterranean rodents. The first description of their chromosome variability was published almost 40 years ago. Studying the G-band structure of chromosomes in numerous individuals revealed subsequent homologous, step-by-step, Robertsonian translocations, which changed diploid numbers from 54 to 30. Here we used a molecular cytogenetic strategy which demonstrates that chromosomal translocations are not always homologous; consequently, karyotypes with the same diploid number can carry different combinations of metacentrics. We further showed that at least three chromosomal forms with 2n = 34 and distinct metacentrics inhabit the Pamir-Alay mountains. Each of these forms independently hybridized with E. tancrei , 2n = 54, forming separate hybrid zones. The chromosomal variations correlate slightly with geographic barriers. Additionally, we confirmed that the emergence of partial or monobrachial homology appeared to be a strong barrier for hybridization in nature, in contradistinction to experiments which we reported earlier. We discuss the possibility of whole arm reciprocal translocations for mole voles. Our findings suggest that chromosomal translocations lead to diversification and speciation.

Researchers have assembled thousands of eukaryotic genomes using Illumina reads, but traditional mate-pair libraries cannot span all repetitive elements, resulting in highly fragmented assemblies. However, both chromosome conformation capture techniques, such as Hi-C and Dovetail Genomics Chicago libraries and long-read sequencing, such as Pacific Biosciences and Oxford Nanopore, help span and resolve repetitive regions and therefore improve genome assemblies. One important livestock species of arid regions that does not have a high-quality contiguous reference genome is the dromedary (Camelus dromedarius). Draft genomes exist but are highly fragmented, and a high-quality reference genome is needed to understand adaptation to desert environments and artificial selection during domestication. Dromedaries are among the last livestock species to have been domesticated, and together with wild and domestic Bactrian camels, they are the only representatives of the Camelini tribe, which highlights their evolutionary significance. Here we describe our efforts to improve the North African dromedary genome. We used Chicago and Hi-C sequencing libraries from Dovetail Genomics to resolve the order of previously assembled contigs, producing almost chromosome-level scaffolds. Remaining gaps were filled with Pacific Biosciences long reads, and then scaffolds were comparatively mapped to chromosomes. Long reads added 99.32 Mbp to the total length of the new assembly. Dovetail Chicago and Hi-C libraries increased the longest scaffold over 12-fold, from 9.71 Mbp to 124.99 Mbp and the scaffold N50 over 50-fold, from 1.48 Mbp to 75.02 Mbp. We demonstrate that Illumina de novo assemblies can be substantially upgraded by combining chromosome conformation capture and long-read sequencing., (© 2019 The Authors. Molecular Ecology Resources Published by John Wiley & Sons Ltd.)

Evolutionary history and taxonomic position for cryptic species may be clarified by using molecular and cytogenetic methods. The subterranean rodent, the Alay mole vole Ellobiusalaicus Vorontsov et al., 1969 is one of three sibling species constituting the subgenus Ellobius Fischer, 1814, all of which lost the Y chromosome and obtained isomorphic XX sex chromosomes in both males and females. E.alaicus is evaluated by IUCN as a data deficient species because their distribution, biology, and genetics are almost unknown. We revealed specific karyotypic variability (2n = 52-48) in E.alaicus due to different Robertsonian translocations (Rbs). Two variants of hybrids (2n = 53, different Rbs) with E.tancrei Blasius, 1884 were found at the Northern slopes of the Alay Ridge and in the Naryn district, Kyrgyzstan. We described the sudden change in chromosome numbers from 2n = 50 to 48 and specific karyotype structure for mole voles, which inhabit the entrance to the Alay Valley (Tajikistan), and revealed their affiliation as E.alaicus by cytochrome b and fragments of nuclear XIST and Rspo1 genes sequencing. To date, it is possible to expand the range of E.alaicus from the Alay Valley (South Kyrgyzstan) up to the Ferghana Ridge and the Naryn Basin, Tien Shan at the north-east and to the Pamir-Alay Mountains (Tajikistan) at the west. The closeness of E.tancrei and E.alaicus is supported, whereas specific chromosome and molecular changes, as well as geographic distribution, verified the species status for E.alaicus . The case of Ellobius species accented an unevenness in rates of chromosome and nucleotide changes along with morphological similarity, which is emblematic for cryptic species.

The sterlet (Acipenser ruthenus Linnaeus, 1758) is a relatively small sturgeon widely distributed in Eurasian rivers from the Danube to the Yenisei. During the twentieth century, all wild sterlet populations have declined due to anthropogenic factors including: overfishing, poaching, construction of dams, and pollution. Despite the necessity of characterization both wild and captive stocks, few studies of population genetics have been performed thus far. Here we studied the genetic diversity and geographic structure of sterlet populations across the eastern range - Ob-Irtysh and Yenisei basins - by sequencing a 628-bp fragment of mitochondrial DNA control region. We identified 98 new haplotypes, delineated 12 haplogroups and estimated the time of basal haplogroup divergence within the species as over 8 million years ago. Our data suggest that Ob-Irtysh and Yenisei populations are isolated from each other and much lower genetic diversity is present in the Yenisei population than in the Ob-Irtysh population. Our data imply that sterlet populations in Siberian rivers underwent bottleneck or fragmentation, followed by subsequent population expansion. The data obtained here are important for sterlet population monitoring and restocking management.

Remarkably stable genomic chromosome elements (evolutionary conserved segments or syntenies) are the basis of large-scale chromosome architecture in vertebrate species. However, these syntenic elements harbour evolutionary important changes through intrachromosomal rearrangements such as inversions and centromere repositioning. Here, using FISH with a set of 20 region-specific probes on a wide array of 28 species, we analyzed evolution of three conserved syntenic regions of the Arvicolinae ancestral karyotype. Inside these syntenies we uncovered multiple, previously cryptic intrachromosomal rearrangements. Although in each of the three conserved blocks we found inversions and centromere repositions, the blocks experienced different types of rearrangements. In two syntenies centromere repositioning predominated, while in the third region, paracentric inversions were more frequent, whereas pericentric inversions were not detected. We found that some of the intrachromosomal rearrangements, mainly paracentric inversions, were synapomorphic for whole arvicoline genera or tribes: genera Alexandromys and Microtus, tribes Ellobini and Myodini. We hypothesize that intrachromosomal rearrangements within conserved syntenic blocks are a major evolutionary force modulating genome architecture in species-rich and rapidly-evolving rodent taxa. Inversions and centromere repositioning may impact speciation and provide a potential link between genome evolution, speciation, and biogeography.

Strains of red fox (Vulpes vulpes) with markedly different behavioural phenotypes have been developed in the famous long-term selective breeding programme known as the Russian farm-fox experiment. Here we sequenced and assembled the red fox genome and re-sequenced a subset of foxes from the tame, aggressive and conventional farm-bred populations to identify genomic regions associated with the response to selection for behaviour. Analysis of the re-sequenced genomes identified 103 regions with either significantly decreased heterozygosity in one of the three populations or increased divergence between the populations. A strong positional candidate gene for tame behaviour was highlighted: SorCS1, which encodes the main trafficking protein for AMPA glutamate receptors and neurexins and suggests a role for synaptic plasticity in fox domestication. Other regions identified as likely to have been under selection in foxes include genes implicated in human neurological disorders, mouse behaviour and dog domestication. The fox represents a powerful model for the genetic analysis of affiliative and aggressive behaviours that can benefit genetic studies of behaviour in dogs and other mammals, including humans.

In the version of this Article originally published, there were some errors in the affiliations: Stephen J. O'Brien's affiliations were incorrectly listed as 8,9; they should have been 7,9. Affiliation 3 was incorrectly named the Institute of Cytology and Genetics of the Russian Academy of Sciences; it should have read Institute of Cytology and Genetics of the Siberian Branch of the Russian Academy of Sciences. Affiliation 4 was incorrectly named the Institute of Molecular and Cell Biology of the Russian Academy of Sciences; it should have read Institute of Molecular and Cellular Biology of the Siberian Branch of the Russian Academy of Sciences. These have now been corrected.

Hybrid sterility is an important step in the speciation process. Hybrids between dwarf hamsters Phodopus sungorus and P. campbelli provide a good model for studies in cytological and genetic mechanisms of hybrid sterility. Previous studies in hybrids detected multiple abnormalities of spermatogenesis and a high frequency of dissociation between the X and Y chromosomes at the meiotic prophase. In this study, we found that the autosomes of the hybrid males and females underwent paring and recombination as normally as their parental forms did. The male hybrids showed a significantly higher frequency of asynapsis and recombination failure between the heterochromatic arms of the X and Y chromosomes than the males of the parental species. Female hybrids as well as the females of the parental species demonstrated a high incidence of centromere misalignment at the XX bivalent and partial asynapsis of the ends of its heterochromatic arms. In all three karyotypes, recombination was completely suppressed in the heterochromatic arm of the X chromosome, where the pseudoautosomal region is located. We propose that this recombination pattern speeds up divergence of the X- and Y-linked pseudoautosomal regions between the parental species and results in their incompatibility in the male hybrids.

Background: Gray whale, Eschrichtius robustus (E. robustus), is a single member of the family Eschrichtiidae, which is considered to be the most primitive in the class Cetacea. Gray whale is often described as a "living fossil". It is adapted to extreme marine conditions and has a high life expectancy (77 years). The assembly of a gray whale genome and transcriptome will allow to carry out further studies of whale evolution, longevity, and resistance to extreme environment., Results: In this work, we report the first de novo assembly and primary analysis of the E. robustus genome and transcriptome based on kidney and liver samples. The presented draft genome assembly is complete by 55% in terms of a total genome length, but only by 24% in terms of the BUSCO complete gene groups, although 10,895 genes were identified. Transcriptome annotation and comparison with other whale species revealed robust expression of DNA repair and hypoxia-response genes, which is expected for whales., Conclusions: This preliminary study of the gray whale genome and transcriptome provides new data to better understand the whale evolution and the mechanisms of their adaptation to the hypoxic conditions.

Several whole genome duplication (WGD) events followed by rediploidization took place in the evolutionary history of vertebrates. Acipenserids represent a convenient model group for investigation of the consequences of WGD as their representatives underwent additional WGD events in different lineages resulting in ploidy level variation between species, and these processes are still ongoing. Earlier, we obtained a set of sterlet (Acipenser ruthenus) chromosome-specific libraries by microdissection and revealed that they painted two or four pairs of whole sterlet chromosomes, as well as additional chromosomal regions, depending on rediploidization status and chromosomal rearrangements after genome duplication. In this study, we employed next generation sequencing to estimate the content of libraries derived from different paralogous chromosomes of sterlet. For this purpose, we aligned the obtained reads to the spotted gar (Lepisosteus oculatus) reference genome to reveal syntenic regions between these two species having diverged 360 Mya. We also showed that the approach is effective for synteny prediction at various evolutionary distances and allows one to clearly distinguish paralogous chromosomes in polyploid genomes. We postulated that after the acipenserid-specific WGD sterlet karyotype underwent multiple interchromosomal rearrangements, but different chromosomes were involved in this process unequally., Competing Interests: The authors declare no conflict of interest.

It has long been hypothesized that chromosomal rearrangements play a central role in different evolutionary processes, particularly in speciation and adaptation. Interchromosomal rearrangements have been extensively mapped using chromosome painting. However, intrachromosomal rearrangements have only been described using molecular cytogenetics in a limited number of mammals, including a few rodent species. This situation is unfortunate because intrachromosomal rearrangements are more abundant than interchromosomal rearrangements and probably contain essential phylogenomic information. Significant progress in the detection of intrachromosomal rearrangement is now possible, due to recent advances in molecular biology and bioinformatics. We investigated the level of intrachromosomal rearrangement in the Arvicolinae subfamily, a species-rich taxon characterized by very high rate of karyotype evolution. We made a set of region specific probes by microdissection for a single syntenic region represented by the p-arm of chromosome 1 of Alexandromys oeconomus , and hybridized the probes onto the chromosomes of four arvicolines ( Microtus agrestis , Microtus arvalis , Myodes rutilus , and Dicrostonyx torquatus ). These experiments allowed us to show the intrachromosomal rearrangements in the subfamily at a significantly higher level of resolution than previously described. We found a number of paracentric inversions in the karyotypes of M. agrestis and M. rutilus , as well as multiple inversions and a centromere shift in the karyotype of M. arvalis . We propose that during karyotype evolution, arvicolines underwent a significant number of complex intrachromosomal rearrangements that were not previously detected., Competing Interests: The authors declare no conflict of interest.

Sokolov's dwarf hamster (Cricetulus sokolovi) is the least studied representative of the striped hamsters (Cricetulus barabensis species group), the taxonomy of which remains controversial. The species was described based on chromosome morphology, but neither the details of the karyotype nor the phylogenetic relationships with other Cricetulus are known. In the present study, the karyotype of C. sokolovi was examined using cross-species chromosome painting. Molecular and cytogenetic data were employed to determine the phylogenetic position of Sokolov's hamster and to analyze the potential pathways of chromosome evolution in Cricetulus. Both the chromosome and molecular data support the species status of Sokolov's hamster. Phylogenetic analysis of the CYTB data placed C. sokolovi as sister to all other striped hamsters (sequence divergence of 8.1%). FISH data revealed that the karyotype of C. sokolovi is highly rearranged, with the most parsimonious scenario of its origin implying at least 4 robertsonian events and a centromere shift. Comparative cytogenetic data on Cricetinae suggest that their evolutionary history includes both periods of chromosomal conservatism and episodes of rapid chromosomal change., (© 2017 S. Karger AG, Basel.)

Acipenseriformes represent a phylogenetically basal clade of ray-finned fish characterized by unusual genomic traits, including paleopolyploid states of extant genomes with high chromosome numbers and slow rates of molecular evolution. Despite a high interest in this fish group, only a limited number of studies have been accomplished on the isolation and characterization of repetitive DNA, karyotype standardization is not yet complete, and sex chromosomes are still to be identified. Here, we applied next-generation sequencing and cluster analysis to characterize major fractions of sterlet (Acipenser ruthenus) repetitive DNA. Using FISH, we mapped 16 tandemly arranged sequences on sterlet chromosomes and found them to be unevenly distributed in the genome with a tendency to cluster in particular regions. Some of the satellite DNAs might be used as specific markers to identify individual chromosomes and their paralogs, resulting in the unequivocal identification of at least 18 chromosome pairs. Our results provide an insight into the characteristic genomic distribution of the most common sterlet repetitive sequences. Biased accumulation of repetitive DNAs in particular chromosomes makes them especially interesting for further search for cryptic sex chromosomes. Future studies of these sequences in other acipenserid species will provide new perspectives regarding the evolution of repetitive DNA within the genomes of this fish order., (© 2017 S. Karger AG, Basel.)

The generic status of Lasiopodomys and its division into subgenera Lasiopodomys (L. mandarinus, L. brandtii) and Stenocranius (L. gregalis, L. raddei) are not generally accepted because of contradictions between the morphological and molecular data. To obtain cytogenetic evidence for the Lasiopodomys genus and its subgenera and to test the autosome to sex chromosome translocation hypothesis of sex chromosome complex origin in L. mandarinus proposed previously, we hybridized chromosome painting probes from the field vole (Microtus agrestis, MAG) and the Arctic lemming (Dicrostonyx torquatus, DTO) onto the metaphases of a female Mandarin vole (L. mandarinus, 2n = 47) and a male Brandt's vole (L. brandtii, 2n = 34). In addition, we hybridized Arctic lemming painting probes onto chromosomes of a female narrow-headed vole (L. gregalis, 2n = 36). Cross-species painting revealed three cytogenetic signatures (MAG12/18, 17a/19, and 22/24) that could validate the genus Lasiopodomys and indicate the evolutionary affinity of L. gregalis to the genus. Moreover, all three species retained the associations MAG1bc/17b and 2/8a detected previously in karyotypes of all arvicolins studied. The associations MAG2a/8a/19b, 8b/21, 9b/23, 11/13b, 12b/18, 17a/19a, and 5 fissions of ancestral segments appear to be characteristic for the subgenus Lasiopodomys. We also validated the autosome to sex chromosome translocation hypothesis on the origin of complex sex chromosomes in L. mandarinus. Two translocations of autosomes onto the ancestral X chromosome in L. mandarinus led to a complex of neo-X1, neo-X2, and neo-X3 elements. Our results demonstrate that genus Lasiopodomys represents a striking example of rapid chromosome evolution involving both autosomes and sex chromosomes. Multiple reshuffling events including Robertsonian fusions, chromosomal fissions, inversions and heterochromatin expansion have led to the formation of modern species karyotypes in a very short time, about 2.4 MY., Competing Interests: The authors have declared that no competing interests exist.

The subfamily Arvicolinae consists of a great number of species with highly diversified karyotypes. In spite of the wide use of arvicolines in biological and medicine studies, the data on their karyotype structures are limited. Here, we made a set of painting probes from flow-sorted chromosomes of a male Palearctic collared lemming (Dicrostonyx torquatus, DTO). Together with the sets of painting probes made previously from the field vole (Microtus agrestis, MAG) and golden hamster (Mesocricetus auratus, MAU), we carried out a reciprocal chromosome painting between these three species. The three sets of probes were further hybridized onto the chromosomes of the Eurasian water vole (Arvicola amphibius) and northern red-backed vole (Myodes rutilus). We defined the diploid chromosome number in D. torquatus karyotype as 2n = 45 + Bs and showed that the system of sex chromosomes is X1X2Y1. The probes developed here provide a genomic tool-kit, which will help to investigate the evolutionary biology of the Arvicolinae rodents. Our results show that the syntenic association MAG1/17 is present not only in Arvicolinae but also in some species of Cricetinae; and thus, should not be considered as a cytogenetic signature for Arvicolinae. Although cytogenetic signature markers for the genera have not yet been found, our data provides insight into the likely ancestral karyotype of Arvicolinae. We conclude that the karyotypes of modern voles could have evolved from a common ancestral arvicoline karyotype (AAK) with 2n = 56 mainly by centric fusions and fissions.

Background: Acipenseriformes take a basal position among Actinopteri and demonstrate a striking ploidy variation among species. The sterlet (Acipenser ruthenus, Linnaeus, 1758; ARUT) is a diploid 120-chromosomal sturgeon distributed in Eurasian rivers from Danube to Enisey. Despite a high commercial value and a rapid population decline in the wild, many genomic characteristics of sterlet (as well as many other sturgeon species) have not been studied., Results: Cell lines from different tissues of 12 sterlet specimens from Siberian populations were established following an optimized protocol. Conventional cytogenetic studies supplemented with molecular cytogenetic investigations on obtained fibroblast cell lines allowed a detailed description of sterlet karyotype and a precise localization of 18S/28S and 5S ribosomal clusters. Localization of sturgeon specific HindIII repetitive elements revealed an increased concentration in the pericentromeric region of the acrocentric ARUT14, while the total sterlet repetitive DNA fraction (C0t30) produced bright signals on subtelomeric segments of small chromosomal elements. Chromosome and region specific probes ARUT1p, 5, 6, 7, 8 as well as 14 anonymous small sized chromosomes (probes A-N) generated by microdissection were applied in chromosome painting experiments. According to hybridization patterns all painting probes were classified into two major groups: the first group (ARUT5, 6, 8 as well as microchromosome specific probes C, E, F, G, H, and I) painted only a single region each on sterlet metaphases, while probes of the second group (ARUT1p, 7 as well as microchromosome derived probes A, B, D, J, K, M, and N) marked two genomic segments each on different chromosomes. Similar results were obtained on male and female metaphases., Conclusions: The sterlet genome represents a complex mosaic structure and consists of diploid and tetraploid chromosome segments. This may be regarded as a transition stage from paleotetraploid (functional diploid) to diploid genome condition. Molecular cytogenetic and genomic studies of other 120- and 240-chromosomal sturgeons are needed to reconstruct genome evolution of this vertebrate group.

The domesticated guinea pig, Cavia porcellus (Hystricomorpha, Rodentia), is an important laboratory species and a model for a number of human diseases. Nevertheless, genomic tools for this species are lacking; even its karyotype is poorly characterized. The guinea pig belongs to Hystricomorpha, a widespread and important group of rodents; so far the chromosomes of guinea pigs have not been compared with that of other hystricomorph species or with any other mammals. We generated full sets of chromosome-specific painting probes for the guinea pig by flow sorting and microdissection, and for the first time, mapped the chromosomal homologies between guinea pig and human by reciprocal chromosome painting. Our data demonstrate that the guinea pig karyotype has undergone extensive rearrangements: 78 synteny-conserved human autosomal segments were delimited in the guinea pig genome. The high rate of genome evolution in the guinea pig may explain why the HSA7/16 and HSA16/19 associations presumed ancestral for eutherians and the three syntenic associations (HSA1/10, 3/19, and 9/11) considered ancestral for rodents were not found in C. porcellus. The comparative chromosome map presented here is a starting point for further development of physical and genetic maps of the guinea pig as well as an aid for genome assembly assignment to specific chromosomes. Furthermore, the comparative mapping will allow a transfer of gene map data from other species. The probes developed here provide a genomic toolkit, which will make the guinea pig a key species to unravel the evolutionary biology of the Hystricomorph rodents.

The karyotypic relationships of skunks (Mephitidae) with other major clades of carnivores are not yet established. Here, multi-directional chromosome painting was used to reveal the karyological relationships among skunks and between Mephitidae (skunks) and Procyonidae (raccoons). Representative species from three genera of Mephitidae (Mephitis mephitis, 2n = 50; Mephitis macroura, 2n = 50; Conepatus leuconotus, 2n = 46; Spilogale gracilis, 2n = 60) and one species of Procyonidae (Procyon lotor, 2n = 38) were studied. Chromosomal homology was mapped by hybridization of five sets of whole-chromosome paints derived from stone marten (Martes foina, 2n = 38), cat, skunks (M. mephitis; M. macroura) and human. The karyotype of the raccoon is highly conserved and identical to the hypothetical ancestral musteloid karyotype, suggesting that procyonids have a particular importance for establishing the karyological evolution within the caniforms. Ten fission events and five fusion events are necessary to generate the ancestral skunk karyotype from the ancestral carnivore karyotype. Our results show that Mephitidae joins Canidae and Ursidae as the third family of carnivores that are characterized by a high rate of karyotype evolution. Shared derived chromosomal fusion of stone marten chromosomes 6 and 14 phylogenetically links the American hog-nosed skunk and eastern spotted skunk.

The cerebellar interposed nuclei (IN) are critical components of a neural network that controls the expression of classically conditioned eyeblinks. The IN receive 2 major inputs: the massive, gamma-aminobutyric acid (GABA)-mediated input from the Purkinje cells of the cerebellar cortex and the relatively weaker, glutamate-mediated input from collaterals of mossy and climbing fiber cerebellar afferent systems. To elucidate the role of IN glutamate neurotransmission in conditioned response (CR) expression, effects of blocking fast glutamatergic neurotransmission in the IN with gamma-d-glutamylglycine (DGG) on the expression of conditioned eyeblinks and on cerebellar nuclear neuronal activity were examined. Surprisingly, blocking fast glutamate receptors in the IN did not abolish CRs. DGG decreased CR incidence and slightly increased CR latency. In contrast, identical amounts of DGG applied to the cerebellar cortex abolished CRs. Similar to the behavioral effects, DGG had unexpectedly mild effects on IN neurons. At the population level, the baseline firing frequency of IN cells was not affected. After DGG injections, the incidence of excitatory modulation of cell activity in the interstimulus interval decreased but was not abolished. A combined block of fast glutamate and GABA(A) neurotransmission using a mixture of DGG and picrotoxin dramatically reduced CR incidence, increased the firing frequency of all cell types, and virtually abolished all modulation of neuronal activity. These results indicate that fast glutamate neurotransmission in the IN plays only an accessory role both in the expression of behavioral CRs and in the generation of associated neuronal activity in the IN.

C-banding and fluorescence in situ hybridization (FISH) document the distribution of constitutive heterochromatin and six highly repeated DNA families (MSAT2570, MSAT21, MSAT160, MS2, MS4 and STR47) in the chromosomes of nine species of Microtus (M. chrotorrhinus, M. rossiaemeridionalis, M. arvalis, M. ilaeus, M. transcaspicus, M. cabrerae, M. pennsylvanicus, M. miurus and M. ochrogaster). Autosomal heterochromatin is largely centromeric and contains different repeated families in different species. Similarly, large C-band positive blocks on the sex chromosomes of four species contain different repeated DNAs. This interspecific variation in the chromosomal distribution and copy number of the repeats suggests that a common ancestor to modern species contained most of the repetitive families, and that descendant species selectively amplified or deleted different repeats on different chromosomes.

Karyotypes of Calomyscus from different regions of Turkmenistan, Iran, and Azerbaijan were studied using chromosome banding (G- and C-banding) and analyses of meiosis in laboratory hybrids. Extensive variation in the diploid number and the number of autosomal arms (FNa) was revealed (2n = 30, FNa = 44; 2n = 32, FNa = 42; 2n = 44, FNa = 46; 2n = 44, FNa = 58; 2n = 37, FNa = 44; 2n = 50, FNa = 50; 2n = 52, FNa = 56). Centric and tandem fusions and heterochromatin changes were identified as the major modes of karyotype evolution in this group. Natural hybrids between individuals with different karyotypes were recorded, and regular chromosome pairing in meiosis was observed in laboratory hybrids. Fluorescence in situ hybridization with a 353-bp BspRI complex tandem repeat indicated that chromosomal repatterning occurred recently within the genus. There is no unequivocal evidence suggesting the role of chromosomal change in the speciation of the populations of Calomyscus examined., (Copyright 2000 S. Karger AG, Basel)

The genes for major ribosomal RNA were localized on chromosomes 5pter-p15, 9q64-qter, and 13q38-qter of the house musk shrew, Suncus murinus (Insectivora, Soricidae) by silver staining of mitotic metaphase and meiotic pachytene spreads and fluorescence in situ hybridization using the human 28S-RNA genes as a probe to mitotic metaphase spreads. The data presented indicate a correlation between sites of in situ hybridization and silver staining. The finding of nuclear materials in mitosis was in a good agreement with observation in meiosis: same chromosomes carried active NORs in both meiotic and mitotic cells.