Publications by authors named "Nikolay B Rubtsov"

20 Publications

  • Page 1 of 1

Two Separate Cases: Complex Chromosomal Abnormality Involving Three Chromosomes and Small Supernumerary Marker Chromosome in Patients with Impaired Reproductive Function.

Genes (Basel) 2020 12 17;11(12). Epub 2020 Dec 17.

Institute of Cytology and Genetics, The Siberian Branch of the Russian Academy of Sciences, 630090 Novosibirsk, Russia.

For medical genetic counseling, estimating the chance of a child being born with chromosome abnormality is crucially important. Cytogenetic diagnostics of parents with a balanced karyotype are a special case. Such chromosome rearrangements cannot be detected with comprehensive chromosome screening. In the current paper, we consider chromosome diagnostics in two cases of chromosome rearrangement in patients with balanced karyotype and provide the results of a detailed analysis of complex chromosomal rearrangement (CCR) involving three chromosomes and a small supernumerary marker chromosome (sSMC) in a patient with impaired reproductive function. The application of fluorescent in situ hybridization, microdissection, and multicolor banding allows for describing analyzed karyotypes in detail. In the case of a CCR, such as the one described here, the probability of gamete formation with a karyotype, showing a balance of chromosome regions, is extremely low. Recommendation for the family in genetic counseling should take into account the obtained result. In the case of an sSMC, it is critically important to identify the original chromosome from which the sSMC has been derived, even if the euchromatin material is absent. Finally, we present our view on the optimal strategy of identifying and describing sSMCs, namely the production of a microdissectional DNA probe from the sSMC combined with a consequent reverse painting.
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http://dx.doi.org/10.3390/genes11121511DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7766715PMC
December 2020

The free-living flatworm .

Evodevo 2020 2;11. Epub 2020 Mar 2.

1European Research Institute for the Biology of Ageing, University of Groningen, University Medical Center Groningen, Antonius Deusinglaan 1, 9713AV Groningen, The Netherlands.

is a free-living flatworm that is emerging as an attractive experimental animal for research on a broad range of biological questions. One feature setting it apart from other flatworms is the successful establishment of transgenesis methods, facilitated by a steady supply of eggs in the form of single-cell zygotes that can be readily manipulated. This, in combination with the transparency of the animal and its small size, creates practical advantages for imaging and fluorescence-activated cell sorting in studies related to stem cell biology and regeneration. can regenerate most of its body parts, including the germline, thanks to the neoblasts, which represent the flatworm stem cell system. Interestingly, neoblasts seem to have a high capacity of cellular maintenance, as can survive up to 210 Gy of γ-irradiation, and partially offset the negative consequence of ageing. As a non-self-fertilizing simultaneous hermaphrodite that reproduces in a sexual manner, is also used to study sexual selection and other evolutionary aspects of sexual reproduction. Work over the past several years has led to the development of molecular resources and tools, including high-quality genome and transcriptome assemblies, transcriptional profiling of the germline and somatic neoblasts, gene knockdown, and in situ hybridization. The increasingly detailed characterization of this animal has also resulted in novel research questions, such as bio-adhesion based on its adhesion-release glands and genome evolution due to its recent whole-genome duplication.
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http://dx.doi.org/10.1186/s13227-020-00150-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7053086PMC
March 2020

Genome and Karyotype Reorganization after Whole Genome Duplication in Free-Living Flatworms of the Genus .

Int J Mol Sci 2020 Jan 20;21(2). Epub 2020 Jan 20.

The Federal Research Center Institute of Cytology and Genetics SB RAS, Lavrentiev ave. 10, 630090 Novosibirsk, Russia.

The genus represents a diverse group of rhabditophoran flatworms with >200 species occurring around the world. Earlier we uncovered karyotype instability linked to hidden polyploidy in both (2 = 8) and its sibling species (2 = 10), prompting interest in the karyotype organization of close relatives. In this study, we investigated chromosome organization in two recently described and closely related species, and , and explored karyotype instability in laboratory lines and cultures of (DV1/10, 2 = 10) and in more detail. We revealed that three of the four studied species are characterized by karyotype instability, while showed a stable 2 = 6 karyotype. Next, we performed comparative cytogenetics of these species using fluorescent in situ hybridization (FISH) with a set of DNA probes (including microdissected DNA probes generated from chromosomes, rDNA, and telomeric DNA). To explore the chromosome organization of the unusual 2 = 9 karyotype discovered in , we then generated chromosome-specific DNA probes for all chromosomes of this species. Similar to and , our findings suggest that arose via whole genome duplication (WGD) followed by considerable chromosome reshuffling. We discuss possible evolutionary scenarios for the emergence and reorganization of the karyotypes of these species and consider their suitability as promising animal models for studying the mechanisms and regularities of karyotype and genome evolution after a recent WGD.
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http://dx.doi.org/10.3390/ijms21020680DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7013459PMC
January 2020

Sequence Composition and Evolution of Mammalian B Chromosomes.

Genes (Basel) 2018 Oct 10;9(10). Epub 2018 Oct 10.

Severtzov Institute of Ecology and Evolution, Russia Academy of Sciences, Leninsky Pr. 33, Moscow 119071, Russia.

B chromosomes (Bs) revealed more than a hundred years ago remain to be some of the most mysterious elements of the eukaryotic genome. Their origin and evolution, DNA composition, transcriptional activity, impact on adaptiveness, behavior in meiosis, and transfer to the next generation require intensive investigations using modern methods. Over the past years, new experimental techniques have been applied and helped us gain a deeper insight into the nature of Bs. Here, we consider mammalian Bs, taking into account data on their DNA sequencing, transcriptional activity, positions in nuclei of somatic and meiotic cells, and impact on genome functioning. Comparative cytogenetics of Bs suggests the existence of different mechanisms of their formation and evolution. Due to the long and complicated evolvement of Bs, the similarity of their morphology could be explained by the similar mechanisms involved in their development while the difference between Bs even of the same origin could appear due to their positioning at different stages of their evolution. A complex analysis of their DNA composition and other features is required to clarify the origin and evolutionary history of Bs in the species studied. The intraspecific diversity of Bs makes this analysis a very important element of B chromosome studies.
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http://dx.doi.org/10.3390/genes9100490DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6211034PMC
October 2018

Low-pass single-chromosome sequencing of human small supernumerary marker chromosomes (sSMCs) and Apodemus B chromosomes.

Chromosoma 2018 09 30;127(3):301-311. Epub 2018 Jan 30.

Institute of Molecular and Cellular Biology Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.

Supernumerary chromosomes sporadically arise in many eukaryotic species as a result of genomic rearrangements. If present in a substantial part of species population, those are called B chromosomes, or Bs. This is the case for 70 mammalian species, most of which are rodents. In humans, the most common types of extra chromosomes, sSMCs (small supernumerary marker chromosomes), are diagnosed in approximately 1 of 2000 postnatal cases. Due to low frequency in population, human sSMCs are not considered B chromosomes. Genetic content of both B-chromosomes and sSMCs in most cases remains understudied. Here, we apply microdissection of single chromosomes with subsequent low-pass sequencing on Ion Torrent PGM and Illumina MiSeq to identify unique and repetitive DNA sequences present in a single human sSMC and several B chromosomes in mice Apodemus flavicollis and Apodemus peninsulae. The pipeline for sequencing data analysis was made available in Galaxy interface as an addition to previously published command-line version. Human sSMC was attributed to the proximal part of chromosome 15 long arm, and breakpoints leading to its formation were located into satellite DNA arrays. Genetic content of Apodemus B chromosomes was species-specific, and minor alterations were observed in both species. Common features of Bs in these Apodemus species were satellite DNA and ERV enrichment, as well as the presence of the vaccinia-related kinase gene Vrk1. Understanding of the non-essential genome elements content provides important insights into genome evolution in general.
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http://dx.doi.org/10.1007/s00412-018-0662-0DOI Listing
September 2018

Chromosome Evolution in the Free-Living Flatworms: First Evidence of Intrachromosomal Rearrangements in Karyotype Evolution of Macrostomum lignano (Platyhelminthes, Macrostomida).

Genes (Basel) 2017 Oct 30;8(11). Epub 2017 Oct 30.

The Federal Research Center Institute of Cytology and Genetics SB RAS, Lavrentiev ave., 10, Novosibirsk 630090, Russia.

The free-living flatworm is a hidden tetraploid. Its genome was formed by a recent whole genome duplication followed by chromosome fusions. Its karyotype (2n = 8) consists of a pair of large chromosomes (MLI1), which contain regions of all other chromosomes, and three pairs of small metacentric chromosomes. Comparison of MLI1 with metacentrics was performed by painting with microdissected DNA probes and fluorescent in situ hybridization of unique DNA fragments. Regions of MLI1 homologous to small metacentrics appeared to be contiguous. Besides the loss of DNA repeat clusters (pericentromeric and telomeric repeats and the 5S rDNA cluster) from MLI1, the difference between small metacentrics MLI2 and MLI4 and regions homologous to them in MLI1 were revealed. Abnormal karyotypes found in the inbred DV1/10 subline were analyzed, and structurally rearranged chromosomes were described with the painting technique, suggesting the mechanism of their origin. The revealed chromosomal rearrangements generate additional diversity, opening the way toward massive loss of duplicated genes from a duplicated genome. Our findings suggest that the karyotype of is in the early stage of genome diploidization after whole genome duplication, and further studies on and closely related species can address many questions about karyotype evolution in animals.
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http://dx.doi.org/10.3390/genes8110298DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5704211PMC
October 2017

Spatial organization of fibroblast and spermatocyte nuclei with different B-chromosome content in Korean field mouse, Apodemus peninsulae (Rodentia, Muridae).

Genome 2017 Oct 21;60(10):815-824. Epub 2017 Jul 21.

a Institute of Cytology and Genetics, Siberian Branch of Russian Academy of Sciences, 630090 Novosibirsk, Russia.

Korean field mouse (Apodemus peninsulae) shows a wide variation in the number of B chromosomes composed of constitutive heterochromatin. For this reason, it provides a good model to study the influence of the number of centromeres and amount of heterochromatin on spatial organization of interphase nuclei. We analyzed the three-dimensional organization of fibroblast and spermatocyte nuclei of the field mice carrying a different number of B chromosomes using laser scanning microscopy and 3D fluorescence in situ hybridization. We detected a co-localization of the B chromosomes with constitutive heterochromatin of the chromosomes of the basic set. We showed a non-random distribution of B chromosomes in the spermatocyte nuclei. Unpaired B chromosomes showed a tendency to occur in the compartment formed by the unpaired part of the XY bivalent.
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http://dx.doi.org/10.1139/gen-2017-0029DOI Listing
October 2017

New insights into the karyotype evolution of the free-living flatworm Macrostomum lignano (Platyhelminthes, Turbellaria).

Sci Rep 2017 07 20;7(1):6066. Epub 2017 Jul 20.

The Federal Research Center Institute of Cytology and Genetics SB RAS, Lavrentiev ave., 10, Novosibirsk, 630090, Russian Federation.

The free-living flatworm Macrostomum lignano is a model organism for evolutionary and developmental biology studies. Recently, an unusual karyotypic diversity was revealed in this species. Specifically, worms are either 'normal' 2n = 8, or they are aneuploid with one or two additional large chromosome(s) (i.e. 2n = 9 or 2n = 10, respectively). Aneuploid worms did not show visible behavioral or morphological abnormalities and were successful in reproduction. In this study, we generated microdissected DNA probes from chromosome 1 (further called MLI1), chromosome 2 (MLI2), and a pair of similar-sized smaller chromosomes (MLI3, MLI4). FISH using these probes revealed that MLI1 consists of contiguous regions homologous to MLI2-MLI4, suggesting that MLI1 arose due to the whole genome duplication and subsequent fusion of one full chromosome set into one large metacentric chromosome. Therefore, one presumably full haploid genome was packed into MLI1, leading to hidden tetraploidy in the M. lignano genome. The study of Macrostomum sp. 8 - a sibling species of M. lignano - revealed that it usually has one additional pair of large chromosomes (2n = 10) showing a high homology to MLI1, thus suggesting hidden hexaploidy in its genome. Possible evolutionary scenarios for the emergence of the M. lignano and Macrostomum sp. 8 genomes are discussed.
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http://dx.doi.org/10.1038/s41598-017-06498-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5519732PMC
July 2017

Evidence for Karyotype Polymorphism in the Free-Living Flatworm, Macrostomum lignano, a Model Organism for Evolutionary and Developmental Biology.

PLoS One 2016 18;11(10):e0164915. Epub 2016 Oct 18.

Institute of Cytology and Genetics SB RAS, Novosibirsk, Russian Federation.

Over the past decade, the free-living flatworm Macrostomum lignano has been successfully used in many areas of biology, including embryology, stem cells, sexual selection, bioadhesion and aging. The increased use of this powerful laboratory model, including the establishment of genomic resources and tools, makes it essential to have a detailed description of the chromosome organization of this species, previously suggested to have a karyotype with 2n = 8 and one pair of large and three pairs of small metacentric chromosomes. We performed cytogenetic analyses for chromosomes of one commonly used inbred line of M. lignano (called DV1) and uncovered unexpected chromosome number variation in the form of aneuploidies of the largest chromosomes. These results prompted us to perform karyotypic studies in individual specimens of this and other lines of M. lignano reared under laboratory conditions, as well as in freshly field-collected specimens from different natural populations. Our analyses revealed a high frequency of aneuploids and in some cases other numerical and structural chromosome abnormalities in laboratory-reared lines of M. lignano, and some cases of aneuploidy were also found in freshly field-collected specimens. Moreover, karyological analyses were performed in specimens of three further species: Macrostomum sp. 8 (a close relative of M. lignano), M. spirale and M. hystrix. Macrostomum sp. 8 showed a karyotype that was similar to that of M. lignano, with tetrasomy for its largest chromosome being the most common karyotype, while the other two species showed a simpler karyotype that is more typical of the genus Macrostomum. These findings suggest that M. lignano and Macrostomum sp. 8 can be used as new models for studying processes of partial genome duplication in genome evolution.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0164915PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5068713PMC
May 2017

Chromosome morphometry in opisthorchiid species (Platyhelminthes, Trematoda).

Parasitol Int 2017 Aug 9;66(4):396-401. Epub 2016 Jul 9.

Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia; Novosibirsk State University, Novosibirsk, Russia.

Few existing studies have dealt with cytogenetics in trematodes, largely due to the attendant technical difficulty of chromosome preparation. We performed a comparative analysis of chromosomes in five opistorchiid species, including Opisthorchis felineus Rivolta, 1884, Opisthorchis viverrini Poirier, 1886, Clonorchis sinensis Cobbold, 1875, Metorchis xanthosomus Creplin 1846, and Metorchis bilis (Braun, 1790) Odening, 1962. For some of these species, no detailed morphometric description of their karyotypes has yet been published; for the karyotype of Metorchis bilis this is the first-ever description. We found that opisthorchiids, like other trematodes, are characterized by karyotypic conservatism (N=6-7) and karyotype asymmetry, although comparison of chromosome morphometric traits did reveal differences between the karyotypes of the species. Moreover, to address certain a methodological issue in trematode chromosome preparation, we analyzed how the source of chromosomal material (partenitae or mature flukes) and the chromosome preparation techniques used (air-drying and cell suspension methods) affected chromosome spreading and size, concluding that the most reliable comparative method involves comparing relative parameters (relative length, arm ratio, centromeric index) of chromosomes prepared using the same technique.
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http://dx.doi.org/10.1016/j.parint.2016.07.004DOI Listing
August 2017

Sex chromosome synapsis and recombination in male guppies.

Zebrafish 2015 Apr 21;12(2):174-80. Epub 2015 Jan 21.

1 Institute of Cytology and Genetics, The Siberian Branch of the Russian Academy of Sciences , Novosibirsk, Russia .

Guppy X and Y chromosomes represent an early stage in sex chromosome divergence. Synapsis and recombination between X and Y chromosomes attract special attention because recombination suppression promotes their differentiation, but previous studies have given contradictory results. Linkage analysis indicated that recombination between X and Y was extremely rare (<10%) and occurred in the medial part of the Y chromosome, while cytological analysis demonstrated regular association between the distal ends of the X and Y at diakinesis. In this study, we examine pairing and recombination between X and Y chromosomes using immunolocalization of MLH1 to mark recombination nodules, and genomic in situ hybridization with a male DNA probe to identify the Y-specific heterochromatic region. Pairing between X and Y is initiated distally. Single crossovers were detected in 87% of XY synaptonemal complexes, most often in the distal region and less frequently in a median position indicating that end-to-end associations between X and Y are chiasmatic. Thus, we suggest that the very low frequency of recombination detected by linkage analysis in a previous study resulted from a lack of informative markers in distal regions.
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http://dx.doi.org/10.1089/zeb.2014.1000DOI Listing
April 2015

DNA probes for FISH analysis of C-negative regions in human chromosomes.

Methods Mol Biol 2013 ;1039:233-42

Institute of Chemical Biology and Fundamental Medicine, Siberian Branch, Russian Academy of Sciences, Novosibirsk, Russian Federation.

Fluorescent in situ hybridization (FISH) is a powerful technology for studying the chromosome organization and aberrations as well as for searching the homology between chromosomal regions in mammals. Currently, FISH is used as a simple, rapid, and reliable technique for analyzing chromosomal rearrangements and assigning chromosomal breakpoints in modern diagnosing of chromosomal pathology. In addition to cloned DNA fragments, the DNA probes produced by sequence-independent polymerase chain reaction are widely used in FISH assays. As a rule, the DNA probes generated from a genomic or chromosomal DNA by whole genome amplification are enriched for repetitive elements and, consequently, efficient FISH analysis requires that repetitive DNA hybridization is suppressed. The linker-adapter polymerase chain reaction (LA-PCR) using the genomic DNA hydrolyzed with HaeIII and RsaI restriction endonucleases allows the repetitive DNA fraction in DNA probe to be decreased and gene-rich DNA to be predominantly amplified. The protocol described here was proposed for production of the DNA probes for enhanced analysis of the C-negative regions in human chromosomes.
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http://dx.doi.org/10.1007/978-1-62703-535-4_19DOI Listing
March 2014

A comparative study of cell-free apoptotic and genomic DNA using FISH and massive parallel sequencing.

Expert Opin Biol Ther 2012 Jun 16;12 Suppl 1:S11-7. Epub 2012 Apr 16.

Institute of Chemical Biology and Fundamental Medicine, Siberian Branch, Russian Academy of Sciences, pr. Lavrentieva 8, Novosibirsk, Russian Federation.

Objective: Study of circulating DNA (cirDNA) generation mechanisms with respect to their influence on the content of cirDNA is very important since it could indicate the best molecular targets for diagnostic applications. Since apoptosis was shown to be one of the main sources of cirDNA, we performed in vitro comparative study of cell-free apoptotic and genomic DNA (gDNA).

Methods: DNA isolated from culture medium of apoptotic human umbilical vein endothelial cells (cm-apoDNA) and the gDNA from the same living cells was analyzed using FISH and sequenced on SOLiD 3 platform.

Results/conclusions: FISH demonstrates overrepresentation of C-positive chromosome regions in cm-apoDNA. SOLiD 3 data show enrichment of cm-apoDNA for Alu repeats: the content of AluJ, AluS and AluY repeats was, respectively, 2.47-fold (standard deviation (SD) 3.6%), 2.45-fold (SD 5.5%) and 2.79-fold (SD 6.1%) higher in cm-apoDNA. By contrast, some of L1 elements were underrepresented in cm-apoDNA: the content of L1MA and L1ME was, respectively, 1.4-fold (SD 22%) and 1.45-fold (SD 9%) lower in cm-apoDNA. In contrast to FISH, these data and the predominant location of Alu repeats in euchromatic regions evidence the non-uniform gDNA degradation during apoptosis leading to the enrichment of cm-apoDNA with coding sequences.
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http://dx.doi.org/10.1517/14712598.2012.670631DOI Listing
June 2012

Comparative cytogenetics of opisthorchid species (Trematoda, Opisthorchiidae).

Parasitol Int 2012 Mar 23;61(1):87-9. Epub 2011 Jul 23.

Institute of Cytology and Genetics Siberian Branch of RAS, Novosibirsk, Russia.

In the present study karyotypes and chromosomes of five species of the family Opisthorchiidae (Opisthorchis felineus (Rivolta, 1884), O. viverrini (Poirier, 1886), Metorchis xanthosomus (Creplin, 1846), M. bilis (Braun, 1893), and Clonorchis sinensis (Cobbold, 1875)) were compared. Karyotypes of O. felineus, M. xanthosomus, M. bilis and C. sinensis consist of two pairs of large meta- and submetacentrics and five pairs of small chromosomes (2n = 14). The karyotype of O. viverrini is 2n = 12, which indicates a fusion of two chromosomes of opisthorchid ancestral karyotype. Analysis of mitotic and meiotic chromosomes was performed by heterologous in situ hybridization of microdissected DNA probes obtained from chromosomes 1 and 2 of O. felineus and chromosomes 1 and 2 of M. xanthosomus. Results of chromosome staining (C- and AgNOR-banding) and FISH of telomeric probes and ribosomal DNA probe on opisthorchid chromosomes were used for chromosome comparison. Data on chromosome number in opisthorchid species were also discussed.
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http://dx.doi.org/10.1016/j.parint.2011.07.006DOI Listing
March 2012

Distribution of repetitive DNA sequences in chromosomes of five opisthorchid species (Trematoda, Opisthorchiidae).

Parasitol Int 2012 Mar 21;61(1):84-6. Epub 2011 Jul 21.

Institute of Cytology and Genetics Siberian Branch of RAS, Novosibirsk, Russia.

Genomes of opisthorchid species are characterized by small size, suggesting a reduced amount of repetitive DNA in their genomes. Distribution of repetitive DNA sequences in the chromosomes of five species of the family Opisthorchiidae (Opisthorchis felineus 2n = 14 (Rivolta, 1884), Opisthorchis viverrini 2n = 12 (Poirier, 1886), Metorchis xanthosomus 2n = 14 (Creplin, 1846), Metorchis bilis 2n = 14 (Braun, 1890), Clonorchis sinensis 2n = 14 (Cobbold, 1875)) was studied with C- and AgNOR-banding, generation of microdissected DNA probes from individual chromosomes and fluorescent in situ hybridization on mitotic and meiotic chromosomes. Small-sized C-bands were discovered in pericentric regions of chromosomes. Ag-NOR staining of opisthorchid chromosomes and FISH with ribosomal DNA probe showed that karyotypes of all studied species were characterized by the only nucleolus organizer region in one of small chromosomes. The generation of DNA probes from chromosomes 1 and 2 of O. felineus and M. xanthosomus was performed with chromosome microdissection followed by DOP-PCR. FISH of obtained microdissected DNA probes on chromosomes of these species revealed chromosome specific DNA repeats in pericentric C-bands. It was also shown that microdissected DNA probes generated from chromosomes could be used as the Whole Chromosome Painting Probes without suppression of repetitive DNA hybridization. Chromosome painting using microdissected chromosome specific DNA probes showed the overall repeat distribution in opisthorchid chromosomes.
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http://dx.doi.org/10.1016/j.parint.2011.06.027DOI Listing
March 2012

Telomeric DNA in chromosomes of five opisthorchid species.

Parasitol Int 2012 Mar 25;61(1):81-3. Epub 2011 Jun 25.

Institute of Cytology and Genetics Siberian Branch of RAS, Novosibirsk, Russia.

The analysis of telomere repeat distribution in chromosomes of five opisthorchid species (Opisthorchis felineus (Rivolta, 1884), Opisthorchis viverrini (Poirier, 1886), Metorchis xanthosomus (Creplin, 1846), Metorchis bilis (Braun, 1890), Clonorchis sinensis (Cobbold, 1875)) was performed with fluorescent in situ hybridization (FISH) of labeled (TTAGGG)n DNA-probe and PNA telomere probe on mitotic and meiotic chromosomes of these species. It was shown that chromosome telomeres of all studied species contain large clusters of (TTAGGG)n telomeric repeats. Interstitial clusters of the (TTAGGG)n repeats have not been revealed in the chromosomes of any studied species even when FISH of PNA telomere probe on pachytene chromosomes was performed. Furthermore interstitial clusters of the (TTAGGG)n repeats have not been detected in the chromosomes of O. viverrini, one of chromosomes of this species is the result of a fusion of two ancestral opisthorchid chromosomes.
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http://dx.doi.org/10.1016/j.parint.2011.06.013DOI Listing
March 2012

Human embryonic stem cell lines isolation, cultivation, and characterization.

In Vitro Cell Dev Biol Anim 2010 Apr 23;46(3-4):284-93. Epub 2010 Feb 23.

Vavilov Institute of General Genetics RAS, Gubkina str., 3, 119991 Moscow, Russia.

A large number of human embryonic stem cell (hESC) lines have been derived worldwide since the first hESC line establishment in 1998. Despite many common characteristics, most important of which is the pluripotency, hESC lines vary significantly in their transcriptional profiles, genetic, and epigenetic state. These differences may arise both from individual genetics of the cell lines and from variations in their handling such as isolation and cultivation. In order to minimize the latter differences, the standardized protocols of cultivation and inter-laboratory comprehensive studies should be performed. In this report, we summarized our experience of derivation and characterization of hESC lines as well as of adaptation of hESCs to novel cultivation protocols. We have successfully derived five hESC lines and characterized them by previously established criteria, including expression of specific markers and the capacity to differentiate both in vitro and in vivo. Four of these lines, namely hESM01-04, were initially derived using mouse fibroblasts as a feeder and currently are maintained under feeder-free, serum-free conditions using mTeSR1 and Matrigel. The fifth line, hESMK05 was derived in feeder-free, serum-free conditions using mTeSR1 and Matrigel. Cell lines retain their pluripotent status and normal karyotype for more than 70 passages and are available to the scientific community.
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http://dx.doi.org/10.1007/s11626-010-9282-6DOI Listing
April 2010

Is there a difference between T- and B-lymphocyte morphology?

J Biomed Opt 2009 Nov-Dec;14(6):064036

Institute of Chemical Kinetics and Combustion, Siberian Branch RAS, Institutskaya 3, Novosibirsk, 630090, Russia.

We characterize T- and B-lymphocytes from several donors, determining cell diameter, ratio of nucleus to cell diameter, and refractive index of the nucleus and cytoplasm for each individual cell. We measure light-scattering profiles with a scanning flow cytometer and invert the signals using a coated sphere as an optical model of the cell and by relying on a global optimization technique. The main difference in morphology of T- and B-lymphocytes is found to be the larger mean diameters of the latter. However, the difference is smaller than the natural biological variability of a single cell. We propose nuclear inhomogeneity as a possible reason for the deviation of measured light-scattering profiles from real lymphocytes from those obtained from the coated sphere model.
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http://dx.doi.org/10.1117/1.3275471DOI Listing
March 2010

DNA content of the B chromosomes in grasshopper Podisma kanoi Storozh. (Orthoptera, Acrididae).

Chromosome Res 2007 10;15(3):315-25. Epub 2007 May 10.

Novosibirsk State University, 630090, Novosibirsk, Russia.

A DNA library derived from the B chromosome of Podisma kanoi was obtained by chromosome microdissection. A total of 153 DNA clones were isolated from the microdissected DNA library. Twenty of them were sequenced. A comparison of B chromosome DNA sequences with sequences of other species from the DDBJ/GenBank/EMBL database ( http://www.ddbj.nig.ac.jp/ ) was performed. Different patterns of signals were observed after FISH with labeled cloned DNA fragments. FISH signals with cloned DNA fragments painted either whole Bs or their different regions. Some clones also gave signals in pericentromeric regions of A chromosomes. Other cloned DNA fragments gave only background-like signals on A and B chromosomes. Comparative FISH analysis of B chromosomes in Podisma kanoi and P. sapporensis with DNA probes derived from the Bs of these species revealed homologous DNA that was confined within pericentromeric and telemetric regions of the B chromosome in P. kanoi. In contrast to the B chromosomes in P. sapporensis containing large regions enriched with rDNA, only a small cluster of rDNA was detected in one of the examined B chromosomes in P. kanoi. The data strongly suggest an independent origin of B chromosomes in two closely related Podisma species.
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http://dx.doi.org/10.1007/s10577-007-1128-zDOI Listing
August 2007

Microdissection and sequence analysis of pericentric heterochromatin from the Drosophila melanogastermutant Suppressor of Underreplication.

Chromosoma 2002 Jul 16;111(2):114-25. Epub 2002 Apr 16.

Department of Zoology and Animal Biology, University of Geneva, 1211 Geneva 4, Switzerland.

In the Suppressor of Underreplication( SuUR) mutant strain of Drosophila melanogaster, the heterochromatin of polytene chromosomes is not underreplicated and, as a consequence, a number of beta-heterochromatic regions acquire a banded structure. The chromocenter does not form in these polytene chromosomes, and heterochromatic regions, normally part of the chromocenter, become accessible to cytological analysis. We generated four genomic DNA libraries from specific heterochromatic regions by microdissection of polytene chromosomes. In situ hybridization of individual libraries onto SuUR polytene chromosomes shows that repetitive DNA sequences spread into the neighboring euchromatic regions. This observation allows the localization of eu-heterochromatin transition zones on polytene chromosomes. We find that genomic scaffolds from the eu-heterochromatin transition zones are enriched in repetitive DNA sequences homologous to those flanking the suppressor of forked gene [ su(f) repeat]. We isolated and sequenced about 300 clones from the heterochromatic DNA libraries obtained. Most of the clones contain repetitive DNA sequences; however, some of the clones have unique DNA sequences shared with parts of unmapped genomic scaffolds. Hybridization of these clones onto SuUR polytene chromosomes allowed us to assign the cytological localizations of the corresponding genomic scaffolds within heterochromatin. Our results demonstrate that the SuUR mutant renders possible the mapping of heterochromatic scaffolds on polytene chromosomes.
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http://dx.doi.org/10.1007/s00412-002-0190-8DOI Listing
July 2002