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Journal articles on the topic 'DNA; Chromosomes'
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1
Csonka, E., I. Cserpan, K. Fodor, G. Hollo, R. Katona, J. Kereso, T. Praznovszky, et al. "Novel generation of human satellite DNA-based artificial chromosomes in mammalian cells." Journal of Cell Science 113, no. 18 (September 15, 2000): 3207–16. http://dx.doi.org/10.1242/jcs.113.18.3207.
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An in vivo approach has been developed for generation of artificial chromosomes, based on the induction of intrinsic, large-scale amplification mechanisms of mammalian cells. Here, we describe the successful generation of prototype human satellite DNA-based artificial chromosomes via amplification-dependent de novo chromosome formations induced by integration of exogenous DNA sequences into the centromeric/rDNA regions of human acrocentric chromosomes. Subclones with mitotically stable de novo chromosomes were established, which allowed the initial characterization and purification of these artificial chromosomes. Because of the low complexity of their DNA content, they may serve as a useful tool to study the structure and function of higher eukaryotic chromosomes. Human satellite DNA-based artificial chromosomes containing amplified satellite DNA, rDNA, and exogenous DNA sequences were heterochromatic, however, they provided a suitable chromosomal environment for the expression of the integrated exogenous genetic material. We demonstrate that induced de novo chromosome formation is a reproducible and effective methodology in generating artificial chromosomes from predictable sequences of different mammalian species. Satellite DNA-based artificial chromosomes formed by induced large-scale amplifications on the short arm of human acrocentric chromosomes may become safe or low risk vectors in gene therapy.
2
Spell, R. M., and C. Holm. "Nature and distribution of chromosomal intertwinings in Saccharomyces cerevisiae." Molecular and Cellular Biology 14, no. 2 (February 1994): 1465–76. http://dx.doi.org/10.1128/mcb.14.2.1465.
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To elucidate yeast chromosome structure and behavior, we examined the breakage of entangled chromosomes in DNA topoisomerase II mutants by hybridization to chromosomal DNA resolved by pulsed-field gel electrophoresis. Our study reveals that large and small chromosomes differ in the nature and distribution of their intertwinings. Probes to large chromosomes (450 kb or larger) detect chromosome breakage, but probes to small chromosomes (380 kb or smaller) reveal no breakage products. Examination of chromosomes with one small arm and one large arm suggests that the two arms behave independently. The acrocentric chromosome XIV breaks only on the long arm, and its preferred region of breakage is approximately 200 kb from the centromere. When the centromere of chromosome XIV is relocated, the preferred region of breakage shifts accordingly. These results suggest that large chromosomes break because they have long arms and small chromosomes do not break because they have small arms. Indeed, a small metacentric chromosome can be made to break if it is rearranged to form a telocentric chromosome with one long arm or a ring with an "infinitely" long arm. These results suggest a model of chromosomal intertwining in which the length of the chromosome arm prevents intertwinings from passively resolving off the end of the arm during chromosome segregation.
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Spell, R. M., and C. Holm. "Nature and distribution of chromosomal intertwinings in Saccharomyces cerevisiae." Molecular and Cellular Biology 14, no. 2 (February 1994): 1465–76. http://dx.doi.org/10.1128/mcb.14.2.1465-1476.1994.
Full textAbstract:
To elucidate yeast chromosome structure and behavior, we examined the breakage of entangled chromosomes in DNA topoisomerase II mutants by hybridization to chromosomal DNA resolved by pulsed-field gel electrophoresis. Our study reveals that large and small chromosomes differ in the nature and distribution of their intertwinings. Probes to large chromosomes (450 kb or larger) detect chromosome breakage, but probes to small chromosomes (380 kb or smaller) reveal no breakage products. Examination of chromosomes with one small arm and one large arm suggests that the two arms behave independently. The acrocentric chromosome XIV breaks only on the long arm, and its preferred region of breakage is approximately 200 kb from the centromere. When the centromere of chromosome XIV is relocated, the preferred region of breakage shifts accordingly. These results suggest that large chromosomes break because they have long arms and small chromosomes do not break because they have small arms. Indeed, a small metacentric chromosome can be made to break if it is rearranged to form a telocentric chromosome with one long arm or a ring with an "infinitely" long arm. These results suggest a model of chromosomal intertwining in which the length of the chromosome arm prevents intertwinings from passively resolving off the end of the arm during chromosome segregation.
4
Zadesenets, K. S., and N. B. Rubtsov. "Regions enriched for DNA repeats in chromosomes of Macrostomum mirumnovem, a species with a recent Whole Genome Duplication." Vavilov Journal of Genetics and Breeding 24, no. 6 (October 28, 2020): 636–42. http://dx.doi.org/10.18699/vj20.657.
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The free-living flatworm Macrostomum mirumnovem is a neopolyploid species whose genome underwent a recent Whole Genome Duplication (WGD). In the result of chromosome fusions of the ancient haploid chromosome set, large metacentric chromosomes were formed. In addition to three pairs of small metacentrics, the current karyotype of M. mirumnovem contains two pairs of large metacentric chromosomes, MMI1 and MMI2. The generation of microdissected DNA libraries enriched for DNA repeats followed by DNA probe preparation and fluorescent in situ hybridization (FISH) were performed. The DNA probes obtained marked chromosome regions enriched for different DNA repeats in the M. mirumnovem chromosomes. The size and localization of these regions varied in different copies of large chromosomes. They varied even in homologous chromosomes, suggesting their divergence due to genome re-diploidization after a WGD. Besides the newly formed chromosome regions enriched for DNA repeats, B chromosomes were found in the karyotypes of the studied specimens of M. mirumnovem. These B chromosomes varied in size and morphology. FISH with microdissected DNA probes revealed that some Bs had a distinct DNA content. FISH could paint differently B chromosomes in different worms and even in the same sample. B chromosomes could carry a bright specific fluorescent signal or could show no fluorescent signal at all. In latter cases, the specific FISH signal could be absent even in the pericentromeric region of the B chromosome. Possible mechanisms of B chromosome formation and their further evolution are discussed. The results obtained indicate an important role that repetitive DNAs play in genome re-diploidization initiating a rapid differentiation of large chromosome copies. Taking together, karyotype peculiarities (a high level of intraspecific karyotypic diversity associated with chromosome number variation, structural chromosomal rearrangements, and the formation of new regions enriched for DNA repeats) and some phenotypic features of M. mirumnovem (small body size, short lifecycle, easy maintenance in the laboratory) make this species a perspective model in the studies of genomic and karyotypic evolution in species passed through a recent WGD event.
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Willhoeft, Ute, Jutta Mueller-Navia, and Gerald Franz. "Analysis of the sex chromosomes of the Mediterranean fruit fly by microdissected DNA probes." Genome 41, no. 1 (February 1, 1998): 74–78. http://dx.doi.org/10.1139/g97-102.
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In the Mediterranean fruit fly, Ceratitis capitata, the sex-determining region maps to the long arm of the Y chromosome. DNA from this region of the Y chromosome and, for comparison, from the tip of the long arm of the X chromosome, was isolated by microdissection and amplified by degenerate oligonucleotide primer PCR (DOP-PCR). FISH of the Y-chromosomal microdissection products medY1-medY5 to mitotic chromosomes revealed hybridization signals on most of the long arm of the Y chromosome, including the male-determining region, and on the long arm of the X chromosome, as well as weaker signals on the autosomes, some of which were located in the heterochromatin next to the centromeres. The X-chromosomal microdissected probe medX1 revealed strong signals on the sex chromosomes and randomly distributed signals on the autosomes. Chromosomal in situ suppression hybridization indicates that the Y chromosome contains considerable amounts of Y-enriched and Y-specific sequences and that X-enriched sequences are present on the long arm of the X chromosome. The microdissected probes medY1, medY2, and medX1 hybridize to the sex chromosomes of two closely related species,Ceratitis rosa and Trirhithrum coffeae.
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Rovatsos, Michail, Juan Alberto Marchal, Eva Giagia-Athanasopoulou, and Antonio Sánchez. "Molecular Composition of Heterochromatin and Its Contribution to Chromosome Variation in the Microtus thomasi/Microtus atticus Species Complex." Genes 12, no. 6 (May 25, 2021): 807. http://dx.doi.org/10.3390/genes12060807.
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The voles of the Microtus thomasi/M. atticus species complex demonstrate a remarkable variability in diploid chromosomal number (2n = 38–44 chromosomes) and sex chromosome morphology. In the current study, we examined by in situ hybridization the topology of four satellite DNA motifs (Msat-160, Mth-Alu900, Mth-Alu2.2, TTAGGG telomeric sequences) and two transposons (LINE, SINE) on the karyotypes of nine chromosome races (i.e., populations with unique cytogenetic traits) of Microtus thomasi, and two chromosomal races of M. atticus. According to the topology of the repetitive DNA motifs, we were able to identify six types of biarmed chromosomes formed from either Robertsonian or/and tandem fusions. In addition, we identified 14 X chromosome variants and 12 Y chromosome variants, and we were able to reconstruct their evolutionary relations, caused mainly by distinct mechanisms of amplification of repetitive DNA elements, including the telomeric sequences. Our study used the model of the Microtus thomasi/M. atticus species complex to explore how repetitive centromeric content can alter from chromosomal rearrangements and can shape the morphology of sex chromosomes, resulting in extensive inter-species cytogenetic variability.
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Scutt, Charles P., Yasuko Kamisugi, Philip M. Gilmartin, and Fukumi Sakai. "Laser isolation of plant sex chromosomes: studies on the DNA composition of the X and Y sex chromosomes of Silene latifolia." Genome 40, no. 5 (October 1, 1997): 705–15. http://dx.doi.org/10.1139/g97-793.
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X and Y sex chromosomes from the dioecious plant Silene latifolia (white campion) were isolated from mitotic metaphase chromosome preparations on polyester membranes. Autosomes were ablated using an argon ion laser microbeam and isolated sex chromosomes were then recovered on excised fragments of polyester membrane. Sex chromosome associated DNA sequences were amplified using the degenerate oligonucleotide primed polymerase chain reaction (DOP–PCR) and pools of DOP–PCR products were used to investigate the genomic organization of the S. latifolia sex chromosomes. The chromosomal locations of cloned sex chromosome repeat sequences were analysed by fluorescence in situ hybridization and data complementary to laser ablation studies were obtained by genomic in situ hybridization. In combination, these studies demonstrate that the X and Y sex chromosomes of S. latifolia are of very similar DNA composition and also that they share a significant repetitive DNA content with the autosomes. The evolution of sex chromosomes in Silene is discussed and compared with that in another dioecious species, Rumex acetosa.Key words: FISH, GISH, laser-microdissection, sex chromosome, Silene latifolia.
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Dvořák, J., H. B. Zhang, R. S. Kota, and M. Lassner. "Organization and evolution of the 5S ribosomal RNA gene family in wheat and related species." Genome 32, no. 6 (December 1, 1989): 1003–16. http://dx.doi.org/10.1139/g89-545.
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Variation in restriction fragments in nullisomic–tetrasomic and ditelosomic lines of Triticum aestivum 'Chinese Spring' and disomic and ditelosomic substitutions of chromosomes of diploid species Lophopyrum elongatum, T. monococcum ssp. aegilopoides, T. tauschii, and T. umbellulatum for 'Chinese Spring' chromosomes were used to identify chromosomal loci of 5S rRNA genes (5S DNA) in wheat and related species. These loci are on wheat chromosome arms 1BS, 1DS, 5AS, 5BS, and tentatively 5DS, T. m. aegilopoides chromosomes 1A and 5A, T. tauschii chromosomes 1D and 5D, and T. umbellulatum chromosome 5U. In diploid L. elongatum a locus was detected on chromosome arm 1ES. In most genomes, the locus on chromosome 1 contains 5S DNA subfamily with short spacers and the locus on chromosome 5 contains 5S DNA subfamily with long spacers. Only a few genomes were found to be potential exceptions to this rule. Concerted evolution of the 5S DNA loci was examined in several genomes. It appeared that homogenization of spacers occurs predominantly within a locus. A scenario of the evolution of polyploid wheats and relationships among diploid species in Triticum are proposed from the observed variation among 5S DNA loci.Key words: Triticum, Lophopyrum, gene mapping, concerted evolution, wheat evolution, DNA methylation.
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Iannucci, Alessio, Marie Altmanová, Claudio Ciofi, Malcolm Ferguson-Smith, Jorge C. Pereira, Ivan Rehák, Roscoe Stanyon, et al. "Isolating Chromosomes of the Komodo Dragon: New Tools for Comparative Mapping and Sequence Assembly." Cytogenetic and Genome Research 157, no. 1-2 (2019): 123–31. http://dx.doi.org/10.1159/000496171.
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We developed new tools to build a high-quality chromosomal map of the Komodo dragon (Varanus komodoensis) available for cross-species phylogenomic analyses. First, we isolated chromosomes by flow sorting and determined the chromosome content of each flow karyotype peak by FISH. We then isolated additional Komodo dragon chromosomes by microdissection and amplified chromosome-specific DNA pools. The chromosome-specific DNA pools can be sequenced, assembled, and mapped by next-generation sequencing technology. The chromosome-specific paint probes can be used to investigate karyotype evolution through cross-species chromosome painting. Overall, the set of chromosome-specific DNA pools of V. komodoensis provides new tools for detailed phylogenomic analyses of Varanidae and squamates in general.
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Gottesdiener, K., J. Garciá-Anoveros, M. G. Lee, and L. H. Van der Ploeg. "Chromosome organization of the protozoan Trypanosoma brucei." Molecular and Cellular Biology 10, no. 11 (November 1990): 6079–83. http://dx.doi.org/10.1128/mcb.10.11.6079.
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The genome of the protozoan Trypanosoma brucei is known to be diploid. Karyotype analysis has, however, failed to identify homologous chromosomes. Having refined the technique for separating trypanosome chromosomes (L. H. T. Van der Ploeg, C. L. Smith, R. I. Polvere, and K. Gottesdiener, Nucleic Acids Res. 17:3217-3227, 1989), we can now provide evidence for the presence of homologous chromosomes. By determining the chromosomal location of different genetic markers, most of the chromosomes (14, excluding the minichromosomes), could be organized into seven chromosome pairs. In most instances, the putative homologs of a pair differed in size by about 20%. Restriction enzyme analysis of chromosome-sized DNA showed that these chromosome pairs contained large stretches of homologous DNA sequences. From these data, we infer that the chromosome pairs represent homologs. The identification of homologous chromosomes gives valuable insight into the organization of the trypanosome genome, will facilitate the genetic analysis of T. brucei, and suggests the presence of haploid gametes.
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Gottesdiener, K., J. Garciá-Anoveros, M. G. Lee, and L. H. Van der Ploeg. "Chromosome organization of the protozoan Trypanosoma brucei." Molecular and Cellular Biology 10, no. 11 (November 1990): 6079–83. http://dx.doi.org/10.1128/mcb.10.11.6079-6083.1990.
Full textAbstract:
The genome of the protozoan Trypanosoma brucei is known to be diploid. Karyotype analysis has, however, failed to identify homologous chromosomes. Having refined the technique for separating trypanosome chromosomes (L. H. T. Van der Ploeg, C. L. Smith, R. I. Polvere, and K. Gottesdiener, Nucleic Acids Res. 17:3217-3227, 1989), we can now provide evidence for the presence of homologous chromosomes. By determining the chromosomal location of different genetic markers, most of the chromosomes (14, excluding the minichromosomes), could be organized into seven chromosome pairs. In most instances, the putative homologs of a pair differed in size by about 20%. Restriction enzyme analysis of chromosome-sized DNA showed that these chromosome pairs contained large stretches of homologous DNA sequences. From these data, we infer that the chromosome pairs represent homologs. The identification of homologous chromosomes gives valuable insight into the organization of the trypanosome genome, will facilitate the genetic analysis of T. brucei, and suggests the presence of haploid gametes.
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Malimpensa, Geovana C., Josiane B. Traldi, Danyelle Toyama, Flávio Henrique-Silva, Marcelo R. Vicari, and Orlando Moreira-Filho. "Chromosomal Mapping of Repeat DNA in Bergiaria westermanni (Pimelodidae, Siluriformes): Localization of 45S rDNA in B Chromosomes." Cytogenetic and Genome Research 154, no. 2 (2018): 99–106. http://dx.doi.org/10.1159/000487652.
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The occurrence of repetitive DNA in autosomes and B chromosomes of Bergiaria westermanni was examined using conventional and molecular cytogenetic techniques. This species exhibited 2n = 56 chromosomes, with intra- and interindividual variation in the number of heterochromatic B chromosomes (from 0 to 4). The 5S rDNA was localized in pairs 1 and 5, and histone probes (H1, H3, and H4) and U2 small nuclear RNA were syntenic with 5S rDNA in pair 5. Histone sequences were also located in chromosome pair 14. The (GATA)n sequence was dispersed throughout the autosomes and B chromosomes, with clusters (microsatellite accumulation) in some chromosome regions. The telomeric probe revealed no signs of chromosomal rearrangements in the genome of B. westermanni. The 45S rDNA sites were detected in the terminal region of pair 27; these sites corresponded to a GC-rich heterochromatin block. In addition, 3 of the 4 B chromosomes also contained 45S rDNA copies. Silver nitrate staining in interphase nuclei provided indirect evidence of the expression of these rRNA genes in B chromosomes, indicating the probable origin of these elements. This report shows plasticity in the chromosomal localization of repeat DNA in B. westermanni and features a discussion of genomic diversification.
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Valadares-Inglis, Maria Cléria, and John F. Peberdy. "Variation in the electrophoretic karyotype of Brazilian strains of Metarhizium anisopliae." Genetics and Molecular Biology 21, no. 1 (March 1998): 11–14. http://dx.doi.org/10.1590/s1415-47571998000100003.
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Pulsed-field gel electrophoresis (PFGE) was used to separate chromosome-sized DNA molecules of four strains of Metarhizium anisopliae from Brazil. Metarhizium anisopliae isolates from Japan have been reported as possessing seven chromosomes. Variation was observed among the Brazilian strains and the chromosomal DNA was resolved into eight bands for strain CG46. Densitometric analysis of PFGE gels suggested that the other three Brazilian strains also possess eight chromosomes, with two chromosomes migrating as doublets under the electrophoretic conditions used. The genome size was estimated as varying between 23.39 to 31.88 Mb, not including possible doublet chromosomes.
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Swedlow, Jason R., Neil Osheroff, Tim Karr, John W. Sedat, and David A. Agard. "The in vivo distribution and dynamics of DNA topoisomerase II in Drosophila embryonic nuclei and chromosomes." Proceedings, annual meeting, Electron Microscopy Society of America 51 (August 1, 1993): 74–75. http://dx.doi.org/10.1017/s0424820100146217.
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DNA topoisomerase II is an ATP-dependent double-stranded DNA strand-passing enzyme that is necessary for full condensation of chromosomes and for complete segregation of sister chromatids at mitosis in vivo and in vitro. Biochemical characterization of chromosomes or nuclei after extraction with high-salt or detergents and DNAse treatment showed that topoisomerase II was a major component of this remnant, termed the chromosome scaffold. The scaffold has been hypothesized to be the structural backbone of the chromosome, so the localization of topoisomerase II to die scaffold suggested that the enzyme might play a structural role in the chromosome. However, topoisomerase II has not been studied in nuclei or chromosomes in vivo. We have monitored the chromosomal distribution of topoisomerase II in vivo during mitosis in the Drosophila embryo. This embryo forms a multi-nucleated syncytial blastoderm early in its developmental cycle. During this time, the embryonic nuclei synchronously progress through 13 mitotic cycles, so this is an ideal system to follow nuclear and chromosomal dynamics.
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Schmid, Michael, and Claus Steinlein. "The Hypermethylated Regions in Avian Chromosomes." Cytogenetic and Genome Research 151, no. 4 (2017): 216–27. http://dx.doi.org/10.1159/000464268.
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Chromosomal locations and amounts of 5-methylcytosine-rich chromosome regions were detected in the karyotypes of 13 bird species by indirect immunofluorescence using a monoclonal anti-5-methylcytosine antibody. These species belong to 7 orders and 10 families of modern (Neognathae) and primitive (Palaeognathae) birds and are characterized by macro- and microchromosomes as well as ZW sex chromosomes. In all 13 species, the hypermethylated chromosome segments are confined to constitutive heterochromatin. The chromosomal locations of hypermethylated DNA regions in the karyotypes are constant and species-specific. There is no general rule with regard to the distribution of these hypermethylated chromosome regions in the genomes of birds. In most instances, hypermethylated segments are located in the centromeric regions of chromosomes, but in the sex chromosomes, these can also be found in telomeric and interstitial postitions. In most of the species studied, the centromeric heterochromatin in many, if not all, of the microchromosomes is hypermethylated. However, in one species, the only detectable hypermethylated heterochromatic regions are located in one pair of macroautosomes and in the Z sex chromosome, but none of the microchromosomes contains visible quantities of 5-methylcytosine. The analysis of 5-methylcytosine-rich chromosome regions can be very helpful for the comparative cytogenetics of closely related species or subspecies. It also reflects the dynamic evolutionary process operating in the highly repetitive DNA of eukaryotic chromosomes.
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Patkin, Eugene L., Maria E. Kustova, and Ekaterina M. Noniashvili. "DNA-strand breaks in chromosomes of early mouse embryos as detected by in situ nick translation and gap filling." Genome 38, no. 2 (April 1, 1995): 381–84. http://dx.doi.org/10.1139/g95-049.
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The nick translation and gap filling procedures, without external addition of nicking enzymes, were performed in situ on fixed chromosomes of mouse preimplantation and postimplantation embryos and of bone marrow in order to detect possible DNA single-strand breaks (nicks and (or) gaps). All chromosome preparations were made using the same technique. Nick translation of chromosomal DNA with DNA polymerase I (Pol I) or gap filling with the Klenow fragment of Pol I in the presence of biotinylated-dUTP, demonstrated a regular absence of label on chromosomes of postimplantation embryos and bone marrow. No difference in sensitivity was found between the holoenzyme and the Klenow fragment. In preimplantation embryos, the chromosome reactivity in nick translation was highest at the blastocyst stage and varied according to cleavage divisions of the zygote.Key words: DNA-strand break, chromosomes of mouse embryos, in situ nick translation.
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Hemmer, Joerg, and Carmen Hauser. "Chromosomal Composition of Aneuploid Clones with Different DNA Contents in Head and Neck Squamous Cell Carcinomas as Determined by Combined Flow Cytometry and FluorescenceIn SituHybridization." Analytical Cellular Pathology 20, no. 4 (2000): 197–203. http://dx.doi.org/10.1155/2000/235942.
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Studies with DNA flow cytometry (FCM) have shown that DNA contents of aneuploid tumour clones vary in a wide range. The aim of this study was to analyse whether homologous chromosomal changes exist despite the individual differences that may be of general relevance for the development of gross aneuploidy in squamous cell carcinomas of the head and neck. Fluorescencein situhybridization (FISH) with 13 centromere‐specific DNA probes was applied to 3 diploid and 11 aneuploid tumours with DNA indices ranging between 0.8 and 2.2. Disomic and monosomic cell populations were prevalent findings in DNA‐diploid tumours. Polysomies were common in aneuploid tumours. Different degrees of aneusomy for identical chromosomes were recurrent features in aneuploid tumours. FISH signal heterogeneity was identified for all chromosomes. The mean number of aneusomic cell populations identified for DNA‐aneuploid tumours ranged between 1.6 for chromosome 17 and 3.1 for chromosome 3. Inconsistencies between FISH and FCM data may indicate that centromere‐specific DNA probes identify gains and losses of marker DNA due to complex karyotypic rearrangements rather than absolute changes in chromosome numbers. Overall, there was no evidence of the critical involvement of particular chromosomes in the development of different DNA contents.
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Nacry, Philippe, Christine Camilleri, Béatrice Courtial, Michel Caboche, and David Bouchez. "Major Chromosomal Rearrangements Induced by T-DNA Transformation in Arabidopsis." Genetics 149, no. 2 (June 1, 1998): 641–50. http://dx.doi.org/10.1093/genetics/149.2.641.
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Abstract We show that major chromosomal rearrangements can occur upon T-DNA transformation of Arabidopsis thaliana. In the ACL4 line, two T-DNA insertion loci were found; one is a tandem T-DNA insert in a head-to-head orientation, and the other is a truncated insert with only the left part of the T-region. The four flanking DNA regions were isolated and located on the Arabidopsis chromosomes; for both inserts, one side of the T-DNA maps to chromosome 2, whereas the other side maps to chromosome 3. Both chromosome 3 flanking regions map to the same location, despite a 1.4-kb deletion at this point, whereas chromosome 2 flanking regions are located 40 cM apart on the bottom arm of chromosome 2. These results strongly suggest a reciprocal translocation between chromosomes 2 and 3, with the breakpoints located at the T-DNA insertion sites. The interchanged fragments roughly correspond to the 20-cM distal ends of both chromosomes. Moreover, a large inversion, spanning 40 cM on the genetic map, occurs on the bottom arm of chromosome 2. This was confirmed by genetic analyses that demonstrated a strong reduction of recombination in the inverted region. Models for T-DNA integration and the consequences for T-DNA tagging are discussed in light of these results.
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Sehnert, Amy J., Brian Rhees, David Comstock, Eileen de Feo, Gabrielle Heilek, John Burke, and Richard P. Rava. "Optimal Detection of Fetal Chromosomal Abnormalities by Massively Parallel DNA Sequencing of Cell-Free Fetal DNA from Maternal Blood." Clinical Chemistry 57, no. 7 (July 1, 2011): 1042–49. http://dx.doi.org/10.1373/clinchem.2011.165910.
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BACKGROUND Massively parallel DNA sequencing of cell-free fetal DNA from maternal blood can detect fetal chromosomal abnormalities. Although existing algorithms focus on the detection of fetal trisomy 21 (T21), these same algorithms have difficulty detecting trisomy 18 (T18). METHODS Blood samples were collected from 1014 patients at 13 US clinic locations before they underwent an invasive prenatal procedure. All samples were processed to plasma, and the DNA extracted from 119 samples underwent massively parallel DNA sequencing. Fifty-three sequenced samples came from women with an abnormal fetal karyotype. To minimize the intra- and interrun sequencing variation, we developed an optimized algorithm by using normalized chromosome values (NCVs) from the sequencing data on a training set of 71 samples with 26 abnormal karyotypes. The classification process was then evaluated on an independent test set of 48 samples with 27 abnormal karyotypes. RESULTS Mapped sites for chromosomes of interest in the sequencing data from the training set were normalized individually by calculating the ratio of the number of sites on the specified chromosome to the number of sites observed on an optimized normalizing chromosome (or chromosome set). Threshold values for trisomy or sex chromosome classification were then established for all chromosomes of interest, and a classification schema was defined. Sequencing of the independent test set led to 100% correct classification of T21 (13 of 13) and T18 (8 of 8) samples. Other chromosomal abnormalities were also identified. CONCLUSION Massively parallel sequencing is capable of detecting multiple fetal chromosomal abnormalities from maternal plasma when an optimized algorithm is used.
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Roth, M., M. Lin, and D. M. Prescott. "Large scale synchronous mating and the study of macronuclear development in Euplotes crassus." Journal of Cell Biology 101, no. 1 (July 1, 1985): 79–84. http://dx.doi.org/10.1083/jcb.101.1.79.
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After conjugation in hypotrichous ciliates, a new macronucleus is produced from a copy of the micronucleus. This transformation involves large-scale reorganization of DNA, with conversion of the chromosomal micronuclear genome into short, gene-sized DNA molecules in the macronucleus. To study directly the changes that occur during this process, we have developed techniques for synchronous mating of large populations of the hypotrichous ciliate Euplotes crassus. Electron microscope studies show that the micronuclear chromosomes are polytenized during the first 20 h of macronuclear development. The polytene chromosomes lack the band-interband organization observed in other hypotrichs and in the Diptera. Polytenization is followed by transectioning of the chromosomes. We isolated DNA at various times of macronuclear development and found that the average molecular weight of the DNA decreases at the time of chromosome transectioning. In addition, we have shown that a small size group of macronuclear DNA molecules (450-550 base pairs) is excised from the chromosomal DNA approximately 10 h later in macronuclear development.
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Schwarzacher, Trude. "DNA, chromosomes, and in situ hybridization." Genome 46, no. 6 (December 1, 2003): 953–62. http://dx.doi.org/10.1139/g03-119.
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In situ hybridization is a powerful and unique technique that correlates molecular information of a DNA sequence with its physical location along chromosomes and genomes. It thus provides valuable information about physical map position of sequences and often is the only means to determine abundance and distribution of repetitive sequences making up the majority of most genomes. Repeated DNA sequences, composed of units of a few to a thousand base pairs in size, occur in blocks (tandem or satellite repeats) or are dispersed (including transposable elements) throughout the genome. They are often the most variable components of a genome, often being species and, occasionally, chromosome specific. Their variability arises through amplification, diversification and dispersion, as well as homogenization and loss; there is a remarkable correlation of molecular sequence features with chromosomal organization including the length of repeat units, their higher order structures, chromosomal locations, and dispersion mechanisms. Our understanding of the structure, function, organization, and evolution of genomes and their evolving repetitive components enabled many new cytogenetic applications to both medicine and agriculture, particularly in diagnosis and plant breeding.Key words: repetitive DNA, genome organization, sequence evolution, telomere, centromere.
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Rudd, M. Katharine, Robert W. Mays, Stuart Schwartz, and Huntington F. Willard. "Human Artificial Chromosomes with Alpha Satellite-Based De Novo Centromeres Show Increased Frequency of Nondisjunction and Anaphase Lag." Molecular and Cellular Biology 23, no. 21 (November 1, 2003): 7689–97. http://dx.doi.org/10.1128/mcb.23.21.7689-7697.2003.
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ABSTRACT Human artificial chromosomes have been used to model requirements for human chromosome segregation and to explore the nature of sequences competent for centromere function. Normal human centromeres require specialized chromatin that consists of alpha satellite DNA complexed with epigenetically modified histones and centromere-specific proteins. While several types of alpha satellite DNA have been used to assemble de novo centromeres in artificial chromosome assays, the extent to which they fully recapitulate normal centromere function has not been explored. Here, we have used two kinds of alpha satellite DNA, DXZ1 (from the X chromosome) and D17Z1 (from chromosome 17), to generate human artificial chromosomes. Although artificial chromosomes are mitotically stable over many months in culture, when we examined their segregation in individual cell divisions using an anaphase assay, artificial chromosomes exhibited more segregation errors than natural human chromosomes (P < 0.001). Naturally occurring, but abnormal small ring chromosomes derived from chromosome 17 and the X chromosome also missegregate more than normal chromosomes, implicating overall chromosome size and/or structure in the fidelity of chromosome segregation. As different artificial chromosomes missegregate over a fivefold range, the data suggest that variable centromeric DNA content and/or epigenetic assembly can influence the mitotic behavior of artificial chromosomes.
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Bríza, J., B. J. Carroll, V. I. Klimyuk, C. M. Thomas, D. A. Jones, and J. D. Jones. "Distribution of unlinked transpositions of a Ds element from a T-DNA locus on tomato chromosome 4." Genetics 141, no. 1 (September 1, 1995): 383–90. http://dx.doi.org/10.1093/genetics/141.1.383.
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Abstract In maize, receptor sites for unlinked transpositions of Activator (Ac) elements are not distributed randomly. To test whether the same is true in tomato, the receptor sites for a Dissociation (Ds) element derived from Ac, were mapped for 26 transpositions unlinked to a donor T-DNA locus on chromosome 4. Four independent transposed Dss mapped to sites on chromosome 4 genetically unlinked to the donor T-DNA, consistent with a preference for transposition to unlinked sites on the same chromosome as opposed to sites on other chromosomes. There was little preference among the nondonor chromosomes, except perhaps for chromosome 2, which carried seven transposed Dss, but these could not be proven to be independent. However, these data, when combined with those from other studies in tomato examining the distribution of transposed Acs or Dss among nondonor chromosomes, suggest there may be absolute preferences for transposition irrespective of the chromosomal location of the donor site. If true, transposition to nondonor chromosomes in tomato would differ from that in maize, where the preference seems to be determined by the spatial arrangement of chromosomes in the interphase nucleus. The tomato lines carrying Ds elements at known locations are available for targeted transposon tagging experiments.
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Winston, Scott, and David E. Pettijohn. "Relaxation of DNA torsional tension in defined domains of bacterial chromosomes in vivo." Canadian Journal of Microbiology 34, no. 4 (April 1, 1988): 522–25. http://dx.doi.org/10.1139/m88-089.
Full textAbstract:
A procedure is described for selectively relaxing the DNA torsional tension in defined regions of the chromosome of living bacterial cells. Regions of the chromosomal DNA labelled with bromodeoxyuridine are selectively nicked by irradiation of the cells with long-wavelength ultraviolet light and then trimethylpsoralen residues are photobound to the chromosome in vivo. It is demonstrated that the rate of photobinding to the bromouridine-labelled parts of the chromosomes declines relative to the unlabelled parts of the same chromosomes as nicks are introduced into the former regions. The maximal difference in photobinding rates is that expected for the difference between relaxed and negatively supercoiled DNA. Analysis of the number of DNA breaks required for minimizing the photobinding rates permits a calculation of the number of domains of supercoiling per Bacillus subtilis chromosome.
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Kubaláková, M., M. Valárik, J. Barto, J. Vrána, J. Cíhalíková, M. Molnár-Láng, and J. Dolezel. "Analysis and sorting of rye (Secale cereale L.) chromosomes using flow cytometry." Genome 46, no. 5 (October 1, 2003): 893–905. http://dx.doi.org/10.1139/g03-054.
Full textAbstract:
Procedures for chromosome analysis and sorting using flow cytometry (flow cytogenetics) were developed for rye (Secale cereale L.). Suspensions of intact chromosomes were prepared by mechanical homogenization of synchronized root tips after mild fixation with formaldehyde. Histograms of relative fluorescence intensity obtained after the analysis of DAPI-stained chromosomes (flow karyotypes) were characterized and the chromosome content of the DNA peaks was determined. Chromosome 1R could be discriminated on a flow karyotype of S. cereale 'Imperial'. The remaining rye chromosomes (2R7R) could be discriminated and sorted from individual wheatrye addition lines. The analysis of lines with reconstructed karyotypes demonstrated a possibility of sorting translocation chromosomes. Supernumerary B chromosomes could be sorted from an experimental rye population and from S. cereale 'Adams'. Flow-sorted chromosomes were identified by fluorescence in situ hybridization (FISH) with probes for various DNA repeats. Large numbers of chromosomes of a single type sorted onto microscopic slides facilitated detection of rarely occurring chromosome variants by FISH with specific probes. PCR with chromosome-specific primers confirmed the identity of sorted fractions and indicated suitability of sorted chromosomes for physical mapping. The possibility to sort large numbers of chromosomes opens a way for the construction of large-insert chromosome-specific DNA libraries in rye.Key words: chromosome isolation, chromosome sorting, fluorescence in situ hybridization, repetitive DNA sequences, wheat-rye addition lines, B chromosomes, physical mapping.
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Setiawan, Agus Budi, Ari Wibowo, Chee How Teo, Shinji Kikuchi, and Takato Koba. "Repetitive DNA sequences accelerate molecular cytogenetic research in plants with small chromosomes." Indonesian Journal of Biotechnology 24, no. 2 (December 5, 2019): 82. http://dx.doi.org/10.22146/ijbiotech.51726.
Full textAbstract:
Repetitive DNA sequences are highly abundant in plant genomes and are favorable probes for chromosome identification in plants. However, it is difficult to conduct studies on the details of metaphase chromosome structures in plants with small chromosomes due to their highly condensed status. Therefore, identification of homologous chromosomes for karyotyping and analyzing chromosome structures is a challenging issue for cytogeneticists without specific probes and precise chromosome stages. In this study, five repetitive DNA probes, i.e., 5S and 45S ribosomal DNAs (rDNAs), melon centromeric sequence (Cmcent), cucumber subtelomeric sequence (Type I), and microsatellite (CT)10 repeats, were used to identify primary constrictions and homologous chromosomes for karyotyping. Four and two loci of 45S rDNA were respectively observed on metaphase and pachytene chromosomes of Abelia × grandiflora. Cmcent was detected on both primary constrictions of melon pachytene and metaphase chromosomes. Furthermore, one pair of 5S rDNA signals were hybridized on melon metaphase chromosomes. Eight and two loci of 45S and 5S rDNA were respectively detected on cucumber chromosomes. Type I and (CT)10 probes were specifically hybridized on subtelomeric and interstitial regions on the chromosomes, respectively. These results suggest that repetitive DNA sequences are versatile probes for chromosome identification in plants with small chromosomes, particularly for karyotyping analyses.
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Moore, Carol Wood, Judith McKoy, Michelle Dardalhon, Darline Davermann, Marcia Martinez, and Dietrich Averbeck. "DNA Damage-Inducible and RAD52-Independent Repair of DNA Double-Strand Breaks in Saccharomyces cerevisiae." Genetics 154, no. 3 (March 1, 2000): 1085–99. http://dx.doi.org/10.1093/genetics/154.3.1085.
Full textAbstract:
Abstract Chromosomal repair was studied in stationary-phase Saccharomyces cerevisiae, including rad52/rad52 mutant strains deficient in repairing double-strand breaks (DSBs) by homologous recombination. Mutant strains suffered more chromosomal fragmentation than RAD52/RAD52 strains after treatments with cobalt-60 γ irradiation or radiomimetic bleomycin, except after high bleomycin doses when chromosomes from rad52/rad52 strains contained fewer DSBs than chromosomes from RAD52/RAD52 strains. DNAs from both genotypes exhibited quick rejoining following γ irradiation and sedimentation in isokinetic alkaline sucrose gradients, but only chromosomes from RAD52/RAD52 strains exhibited slower rejoining (10 min to 4 hr in growth medium). Chromosomal DSBs introduced by γ irradiation and bleomycin were analyzed after pulsed-field gel electrophoresis. After equitoxic damage by both DNA-damaging agents, chromosomes in rad52/rad52 cells were reconstructed under nongrowth conditions [liquid holding (LH)]. Up to 100% of DSBs were eliminated and survival increased in RAD52/RAD52 and rad52/rad52 strains. After low doses, chromosomes were sometimes degraded and reconstructed during LH. Chromosomal reconstruction in rad52/rad52 strains was dose dependent after γ irradiation, but greater after high, rather than low, bleomycin doses with or without LH. These results suggest that a threshold of DSBs is the requisite signal for DNA-damage-inducible repair, and that nonhomologous end-joining repair or another repair function is a dominant mechanism in S. cerevisiae when homologous recombination is impaired.
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Iourov, Ivan Y., Ilia V. Soloviev, Svetlana G. Vorsanova, Viktor V. Monakhov, and Yuri B. Yurov. "An Approach for Quantitative Assessment of Fluorescence In Situ Hybridization (FISH) Signals for Applied Human Molecular Cytogenetics." Journal of Histochemistry & Cytochemistry 53, no. 3 (March 2005): 401–8. http://dx.doi.org/10.1369/jhc.4a6419.2005.
Full textAbstract:
A number of applied molecular cytogenetic studies require the quantitative assessment of fluorescence in situ hybridization (FISH) signals (for example, interphase FISH analysis of aneuploidy by chromosome enumeration DNA probes; analysis of somatic pairing of homologous chromosomes in interphase nuclei; identification of chromosomal heteromorphism after FISH with satellite DNA probes for differentiation of parental origin of homologous chromosome, etc.). We have performed a pilot study to develop a simple technique for quantitative assessment of FISH signals by means of the digital capturing of microscopic images and the intensity measuring of hybridization signals using Scion Image software, commonly used for quantification of electrophoresis gels. We have tested this approach by quantitative analysis of FISH signals after application of chromosome-specific DNA probes for aneuploidy scoring in interphase nuclei in cells of different human tissues. This approach allowed us to exclude or confirm a low-level mosaic form of aneuploidy by quantification of FISH signals (for example, discrimination of pseudo-monosomy and artifact signals due to over-position of hybridization signals). Quantification of FISH signals was also used for analysis of somatic pairing of homologous chromosomes in nuclei of postmortem brain tissues after FISH with “classical” satellite DNA probes for chromosomes 1, 9, and 16. This approach has shown a relatively high efficiency for the quantitative registration of chromosomal heteromorphism due to variations of centromeric alphoid DNA in homologous parental chromosomes. We propose this approach to be efficient and to be considered as a useful tool in addition to visual FISH signal analysis for applied molecular cytogenetic studies.
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Ocampo-Hafalla, Maria, Sofía Muñoz, Catarina P. Samora, and Frank Uhlmann. "Evidence for cohesin sliding along budding yeast chromosomes." Open Biology 6, no. 6 (June 2016): 150178. http://dx.doi.org/10.1098/rsob.150178.
Full textAbstract:
The ring-shaped cohesin complex is thought to topologically hold sister chromatids together from their synthesis in S phase until chromosome segregation in mitosis. How cohesin stably binds to chromosomes for extended periods, without impeding other chromosomal processes that also require access to the DNA, is poorly understood. Budding yeast cohesin is loaded onto DNA by the Scc2–Scc4 cohesin loader at centromeres and promoters of active genes, from where cohesin translocates to more permanent places of residence at transcription termination sites. Here we show that, at the GAL2 and MET17 loci, pre-existing cohesin is pushed downstream along the DNA in response to transcriptional gene activation, apparently without need for intermittent dissociation or reloading. We observe translocation intermediates and find that the distribution of most chromosomal cohesin is shaped by transcription. Our observations support a model in which cohesin is able to slide laterally along chromosomes while maintaining topological contact with DNA. In this way, stable cohesin binding to DNA and enduring sister chromatid cohesion become compatible with simultaneous underlying chromosomal activities, including but maybe not limited to transcription.
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Uno, Yoshinobu, Chizuko Nishida, Chiyo Takagi, Takeshi Igawa, Naoto Ueno, Masayuki Sumida, and Yoichi Matsuda. "Extraordinary Diversity in the Origins of Sex Chromosomes in Anurans Inferred from Comparative Gene Mapping." Cytogenetic and Genome Research 145, no. 3-4 (2015): 218–29. http://dx.doi.org/10.1159/000431211.
Full textAbstract:
Sex determination in frogs (anurans) is genetic and includes both male and female heterogamety. However, the origins of the sex chromosomes and their differentiation processes are poorly known. To investigate diversity in the origins of anuran sex chromosomes, we compared the chromosomal locations of sex-linked genes in 4 species: the African clawed frog (Xenopus laevis), the Western clawed frog (Silurana/X. tropicalis), the Japanese bell-ring frog (Buergeria buergeri), and the Japanese wrinkled frog (Rana rugosa). Comparative mapping data revealed that the sex chromosomes of X. laevis, X. tropicalis and R. rugosa are different chromosome pairs; however, the sex chromosomes of X. tropicalis and B. buergeri are homologous, although this may represent distinct evolutionary origins. We also examined the status of sex chromosomal differentiation in B. buergeri, which possesses heteromorphic ZW sex chromosomes, using comparative genomic hybridization and chromosome painting with DNA probes from the microdissected W chromosome. At least 3 rearrangement events have occurred in the proto-W chromosome: deletion of the nucleolus organizer region and a paracentric inversion followed by amplification of non-W-specific repetitive sequences.
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Engstrom, Mark D., Allan J. Baker, Judith L. Eger, Rudy Boonstra, and Ronald J. Brooks. "Chromosomal and mitochondrial DNA variation in four laboratory populations of collared lemmings (Dicrostonyx)." Canadian Journal of Zoology 71, no. 1 (January 1, 1993): 42–48. http://dx.doi.org/10.1139/z93-007.
Full textAbstract:
Genetic differentiation among populations and speciation in Dicrostonyx is hypothesized to have resulted from either allopatric divergence in glacial refugia during the Wisconsin or sympatric processes uncorrelated with refugial isolation. We examined chromosomal and mitochondrial DNA variation in four laboratory colonies, representing three species, in a preliminary evaluation of these hypotheses. Chromosomal variation is extensive among populations, diploid numbers ranging from 38 to 50. Autosomal variation appears to be due primarily to Robertsonian rearrangements and additions of supernumerary chromosomes, and is geographically unpatterned. Sex chromosome morphology is geographically structured and correlated with proposed southern and northern refugia. Restriction fragment analysis of mitochondrial DNA revealed two ancient, divergent genotypic assemblages, corresponding to geographic distributions of sex chromosomes. Autosomal variation, and any resulting reproductive isolation, probably is recent and uncorrelated with refugial history, whereas divergence of sex chromosomes and disparate mitochondrial assemblages likely predate the Wisconsin.
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Katsumata, M., and C. W. Lo. "Organization of chromosomes in the mouse nucleus: analysis by in situ hybridization." Journal of Cell Science 90, no. 2 (June 1, 1988): 193–99. http://dx.doi.org/10.1242/jcs.90.2.193.
Full textAbstract:
We used in situ hybridization with various biotinylated DNA probes to characterize the distribution of mouse chromosomes in the nucleus. This entailed an examination of plasmid-marked chromosomes from two strains of transgenic mice and also an examination of the distribution of centromeres and total mouse genomic DNA. Our results show that, regardless of the hybridization probes used or the tissue types examined, with sections of mouse tissues embedded in paraffin all chromosomal DNA appeared to be localized to the nuclear periphery. This was observed in tissues fixed with either crosslinking or non-crosslinking fixatives, and submitted to a variety of prehybridization treatments. Further experiments with non-embedded nuclei fractionated from mouse liver revealed a similar distribution of DNA at the telomeric end of chromosome 3, but for centromeric DNA and total mouse genomic DNA, hybridization signals were observed both in the interior and at the periphery of the nucleus. These observations preclude the exclusive localization of chromosomes to the nuclear periphery. However, they indicate that a subset of centromeres are likely to be associated with the nuclear membrane, and that there is such an association at the telomeric end of chromosome 3. Overall, these results are compatible with previous observations of other investigators showing regions of contact between chromosomes and the nuclear membrane.
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Dulz, Thais Aparecida, Carla Andrea Lorscheider, Viviane Demetrio Nascimento, Rafael Bueno Noleto, Orlando Moreira-Filho, Viviane Nogaroto, and Marcelo Ricardo Vicari. "Comparative cytogenetics among Leporinus friderici and Leporellus vittatus populations (Characiformes, Anostomidae): focus on repetitive DNA elements." Comparative Cytogenetics 13, no. 2 (April 5, 2019): 105–20. http://dx.doi.org/10.3897/compcytogen.v13i2.33764.
Full textAbstract:
Anostomidae are a neotropical fish family rich in number of species. Cytogenetically, they show a conserved karyotype with 2n = 54 chromosomes, although they present intraspecific/interspecific variations in the number and chromosomal location of repetitive DNA sequences. The aim of the present study was to perform a comparative description of the karyotypes of two populations of Leporinusfriderici Bloch, 1794 and three populations of Leporellusvittatus Valenciennes, 1850. We used conventional cytogenetic techniques allied to fluorescence in situ hybridization, using 18S ribosomal DNA (rDNA) and 5S rDNA, a general telomere sequence for vertebrates (TTAGGG)n and retrotransposon (RTE) Rex1 probes. The anostomids in all studied populations presented 2n = 54 chromosomes, with a chromosome formula of 32m + 22sm for L.friderici and 28m + 26sm for L.vittatus. Variations in the number and location of the 5S and 18S rDNA chromosomal sites were observed between L.friderici and L.vittatus populations and species. Accumulation of Rex1 was observed in the terminal region of most chromosomes in all populations, and telomere sequences were located just on all ends of the 54 chromosomes in all populations. The intraspecific and intergeneric chromosomal changes occurred in karyotype differentiation, indicating that minor chromosomal rearrangements had present in anostomid species diversification.
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Sharbel, Timothy F., David M. Green, and Andreas Houben. "B-chromosome origin in the endemic New Zealand frog Leiopelma hochstetteri through sex chromosome devolution." Genome 41, no. 1 (February 1, 1998): 14–22. http://dx.doi.org/10.1139/g97-091.
Full textAbstract:
The endemic New Zealand frog Leiopelma hochstetteri has variable numbers of mitotically stable B chromosomes. To assess whether the B chromosomes were derived from the autosome complement, they were isolated by micromanipulation and their DNA amplified by degenerate oligonucleotide primed PCR. Southern hybridizations of B chromosome DNA probes to genomic DNA from males and females characterized by differing numbers of B chromosomes demonstrated that the B chromosomes were derived from the univalent W sex chromosome characteristic of North Island populations. The presence of homologous B chromosome specific sequences from geographically distinct populations indicates a single origin of the B chromosomes. Furthermore, a primitive homology shared by B chromosomes and the W sex chromosome from an ancestral WZ/ZZ karyotype, which is still present in frogs from Great Barrier Island, shows that the B chromosomes originated soon after the univalent W sex chromosome had originated. Sequence analysis revealed that B chromosome DNA is composed of repeat sequences and has the potential to form stable hairpin structures. The molecular dynamics of these structures may reflect an inherent propensity to undergo rapid change in nucleotide sequence and chromosome structure.
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Kawamura, Ryo, Lisa H. Pope, Morten O. Christensen, Mingxuan Sun, Ksenia Terekhova, Fritz Boege, Christian Mielke, Anni H. Andersen, and John F. Marko. "Mitotic chromosomes are constrained by topoisomerase II–sensitive DNA entanglements." Journal of Cell Biology 188, no. 5 (March 1, 2010): 653–63. http://dx.doi.org/10.1083/jcb.200910085.
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We have analyzed the topological organization of chromatin inside mitotic chromosomes. We show that mitotic chromatin is heavily self-entangled through experiments in which topoisomerase (topo) II is observed to reduce mitotic chromosome elastic stiffness. Single chromosomes were relaxed by 35% by exogenously added topo II in a manner that depends on hydrolysable adenosine triphosphate (ATP), whereas an inactive topo II cleavage mutant did not change chromosome stiffness. Moreover, experiments using type I topos produced much smaller relaxation effects than topo II, indicating that chromosome relaxation by topo II is caused by decatenation and/or unknotting of double-stranded DNA. In further experiments in which chromosomes are first exposed to protease to partially release protein constraints on chromatin, ATP alone relaxes mitotic chromosomes. The topo II–specific inhibitor ICRF-187 blocks this effect, indicating that it is caused by endogenous topo II bound to the chromosome. Our experiments show that DNA entanglements act in concert with protein-mediated compaction to fold chromatin into mitotic chromosomes.
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Bauchan, G. R., and M. A. Hossain. "Constitutive heterochromatin DNA polymorphisms in diploid Medicago sativa ssp. falcata." Genome 42, no. 5 (October 1, 1999): 930–35. http://dx.doi.org/10.1139/g99-038.
Full textAbstract:
A Giemsa C-banding technique was used to study the amount and location of constitutive heterochromatin in diploid (2n = 2x = 16) Medicago sativa ssp. falcata (L.) Arcangeli. Most accessions had the standard C-banding pattern with centromeric bands on all the chromosomes and a prominent heterochromatic band at the nucleolar organizer regions (NOR) of the satellited (SAT) chromosomes. However, we observed in various accessions that constitutive heterochromatic C-bands can exist at the telomeric ends of all the chromosomes. Interstitial bands occurred on the short arms of all chromosomes except for chromosome 3 and on the long arms of chromosomes 1, 2, 3, and 6, only. Rearranged chromosomes such as isochromosomes have been observed for the short arms of chromosomes 2 and 6. This is the first report on the existence of C-banding polymorphisms and the detection of putative isochromsomes in the chromosomes of diploid ssp. falcata which could have contributed to the variation observed in cultivated alfalfa.Key words: alfalfa, C-bands, chromosome, isochromosome.
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Kaback, David B., Dianna Barber, Jim Mahon, Jacque Lamb, and Jerome You. "Chromosome Size-Dependent Control of Meiotic Reciprocal Recombination in Saccharomyces cerevisiae: The Role of Crossover Interference." Genetics 152, no. 4 (August 1, 1999): 1475–86. http://dx.doi.org/10.1093/genetics/152.4.1475.
Full textAbstract:
Abstract In the yeast Saccharomyces cerevisiae, small chromosomes undergo meiotic reciprocal recombination (crossing over) at rates (centimorgans per kilobases) greater than those of large chromosomes, and recombination rates respond directly to changes in the total size of a chromosomal DNA molecule. This phenomenon, termed chromosome size-dependent control of meiotic reciprocal recombination, has been suggested to be important for ensuring that homologous chromosomes cross over during meiosis. The mechanism of this regulation was investigated by analyzing recombination in identical genetic intervals present on different size chromosomes. The results indicate that chromosome size-dependent control is due to different amounts of crossover interference. Large chromosomes have high levels of interference while small chromosomes have much lower levels of interference. A model for how crossover interference directly responds to chromosome size is presented. In addition, chromosome size-dependent control was shown to lower the frequency of homologous chromosomes that failed to undergo crossovers, suggesting that this control is an integral part of the mechanism for ensuring meiotic crossing over between homologous chromosomes.
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Perez, Carl, Gary de Jong, Jan Drayer, and Gyula Hadlaczky. "Satellite DNA-based artificial chromosomes – chromosomal vectors." Trends in Biotechnology 18, no. 10 (October 2000): 402–3. http://dx.doi.org/10.1016/s0167-7799(00)01487-6.
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Hardy, Christine D., Nancy J. Crisona, Michael D. Stone, and Nicholas R. Cozzarelli. "Disentangling DNA during replication: a tale of two strands." Philosophical Transactions of the Royal Society of London. Series B: Biological Sciences 359, no. 1441 (January 29, 2004): 39–47. http://dx.doi.org/10.1098/rstb.2003.1363.
Full textAbstract:
The seminal papers by Watson and Crick in 1953 on the structure and function of DNA clearly enunciated the challenge their model presented of how the intertwined strands of DNA are unwound and separated for replication to occur. We first give a historical overview of the major discoveries in the past 50 years that address this challenge. We then describe in more detail the cellular mechanisms responsible for the unlinking of DNA. No single strategy on its own accounts for the complete unlinking of chromosomes required for DNA segregation to proceed. Rather, it is the combined effects of topoisomerase action, chromosome organization and DNA–condensing proteins that allow the successful partitioning of chromosomes into dividing cells. Finally, we propose a model of chromosome structure, consistent with recent findings, that explains how the problem of unlinking is alleviated by the division of chromosomal DNA into manageably sized domains.
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Gatto, Kaleb Pretto, Lucas H. B. Souza, Juliana Nascimento, Pablo Suárez, and Luciana Bolsoni Lourenço. "Comparative mapping of a new repetitive DNA sequence and chromosome region-specific probes unveiling rearrangements in an Amazonian frog complex." Genome 64, no. 9 (September 2021): 857–68. http://dx.doi.org/10.1139/gen-2020-0199.
Full textAbstract:
The frog species Physalaemus ephippifer exists in the Amazonian region and harbors heteromorphic Z and W chromosomes. A genetic lineage closely related to this species was recognized based on its mitochondrial DNA and RADseq-style markers, but its taxonomic status is still unclear and has been referred to as Lineage 1 of “P. cuvieri”. The heteromorphic sex chromosomes found in P. ephippifer are not present in this lineage and which of its chromosome pairs is homologous to the sex chromosomes of P. ephippifer remain to be elucidated as well as the role of such a karyotypic divergence in the evolution of these frogs. Here, we described a new family of repetitive DNA and used its chromosomal sites along with the markers detected by a probe constructed from the microdissected segment of the Z chromosome of P. ephippifer to infer chromosomal homology. We also analyzed an unnamed species that is considered to be the sister group of the clade composed of Lineage 1 of “P. cuvieri” and P. ephippifer. Our results suggest that complex rearrangements involving the chromosomes that were inferred to be homeologous to the sex chromosomes of P. ephippifer have occurred during the divergence of this group of frogs.
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Cuéllar, T., J. Orellana, E. Belhassen, and J. L. Bella. "Chromosomal characterization and physical mapping of the 5S and the 18S-5.8S-25S ribosomal DNA in Helianthus argophyllus, with new data from Helianthus annuus." Genome 42, no. 1 (February 1, 1999): 110–15. http://dx.doi.org/10.1139/g98-112.
Full textAbstract:
The characterization of the mitotic chromosomes of Helianthus argophyllus by means of Feulgen staining, Giemsa C-banding, and the usual DNA sequence-specific fluorochromes allows a chromosomal classification of this species, and shows that two kinds of heterochromatin co-exist equilocally in its chromosomes. One type is confined to the pericentromeric areas of all the chromosomes and the other is associated with the secondary constrictions of the satellite chromosomes. This species is evolutionarily very close to H. annuus with which it is involved in introgression breeding programs. Since these two species show no intra- or interspecific differences with the above treatments, we have used C-banding followed by DAPI, chromomycin A3 or Acridine Orange, and the fluorescent in situ hybridization (FISH) with 5S and 18S-25S ribosomal DNA probes to characterize further the chromosomes of both species in an attempt to find practically applicable chromosomal markers. Our results confirm the heterogeneity of the heterochromatin in these species and show that neither its distribution nor its response to distinct fluorochrome treatments allows better discrimination of the chromosomes within or between the species. On the other hand, the 18S-5.8S-25S rDNA arrays are located in the secondary constrictions of the satellited SM7, SM10, and ST13 pairs and the 5S-rDNA genes are interstitially placed on the short arm of the SM7 and SM11 chromosomes in both species. This permits these chromosomes to be distinguished and provides markers which may be helpful for further physical mapping of DNA sequences in these species.Key words: chromosome banding, sunflower cytogenetics, heterochromatin, ribosomal DNA mapping, FISH.
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Crepaldi, Carolina, and Patricia P. Parise-Maltempi. "Heteromorphic Sex Chromosomes and Their DNA Content in Fish: An Insight through Satellite DNA Accumulation in Megaleporinus elongatus." Cytogenetic and Genome Research 160, no. 1 (2020): 38–46. http://dx.doi.org/10.1159/000506265.
Full textAbstract:
The repetitive DNA content of fish sex chromosomes provides valuable insights into specificities and patterns of their genetic sex determination systems. In this study, we revealed the genomic satellite DNA (satDNA) content of Megaleporinuselongatus, a Neotropical fish species with Z1Z1Z2Z2/Z1W1Z2W2 multiple sex chromosomes, through high-throughput analysis and graph-based clustering, isolating 68 satDNA families. By physically mapping these sequences in female metaphases, we discovered 15 of the most abundant satDNAs clustered in its chromosomes, 9 of which were found exclusively in the highly heterochromatic W1. This heteromorphic sex chromosome showed the highest amount of satDNA accumulations in this species. The second most abundant family, MelSat02-26, shared FISH signals with the NOR-bearing pair in similar patterns and is linked to the multiple sex chromosome system. Our results demonstrate the diverse satDNA content in M. elongatus, especially in its heteromorphic sex chromosome. Additionally, we highlighted the different accumulation patterns and distribution of these sequences across species by physically mapping these satDNAs in other Anostomidae, Megaleporinusmacrocephalus and Leporinusfriderici (a species without differentiated sex chromosomes).
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Gutierrez, Juana, Gael Aleix-Mata, Juan A. Marchal, María Arroyo, Riccardo Castiglia, and Antonio Sánchez. "Molecular Cytogenetic Analysis of Karyotype and Y Chromosome Conservation in Species of the Genus Talpa (Insectivora)." Cytogenetic and Genome Research 160, no. 5 (2020): 264–71. http://dx.doi.org/10.1159/000507836.
Full textAbstract:
The Talpidae family has a highly stable karyotype. Most of the chromosome studies in this mammal group, however, employed classical cytogenetic techniques. Molecular cytogenetic analyses are still scarce and, for example, no repeated DNA sequences have been described to date. In this work, we used sequence analysis, chromosomal mapping of a LINE1 retroelement sequence, as well as chromosome painting with a whole Y chromosome probe of T. occidentalis to compare the karyotypes of 3 species of the genus Talpa (T. occidentalis, T. romana, and T. aquitania). Our results demonstrate that in Talpa genomes LINE1 sequences are widely distributed on all chromosomes but are enriched in pericentromeric C-band-positive regions. In addition, these LINE1 accumulate on the Y chromosomes of the 3 Talpa species regardless of their euchromatic or heterochromatic condition. Chromosome painting shows that the Y chromosomes in these 3 species are highly conserved. Interestingly, they share sequences with heterochromatic blocks on chromosome pairs 14 and 16 and, to a lesser degree, with the pericentromeric regions of other autosomes. Together, our analyses demonstrate that the repetitive DNA content of chromosomes from Talpa species is highly conserved.
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Hausmann, Michael, Gertrud Dudin, Jacob A. Aten, and Rainer Heilig. "Slit Scan Flow Cytometry Of Isolated Chromosomes Following Fluorescence Hybridization: An Approach Of Online Screening For Specific Chromosomes And Chromosome Translocations." Zeitschrift für Naturforschung C 46, no. 5-6 (June 1, 1991): 433–41. http://dx.doi.org/10.1515/znc-1991-5-616.
Full textAbstract:
The recently developed methods of non radioactive in situ hybridization of chromosomes offer new aspects for chromosome analysis. Fluorescent labelling of hybridized chromosomes or chromosomal subregions allows to facilitate considerably the detection of specific chromosomal abnormalities. Formany biomedical applications (e.g. biological dosimetry in the low dose range), a fast scoring for aberrations (e.g. dicentrics or translocations) in required. Here, we present an approach depending on fluorescence in situ hybridization of isolated suspension chromosomes that indicates the feasibility of a rapid screening for specific chromosomes or translocations by slit scan flow cytometry. Chromosomes of a Chinese hamster x human hybrid cell line were hybridized in suspension with biotinylated human genomic DNA . This DNA was decorated with FITC by a double antibody system against biotin. For flow cytometry the chromosomes were stabilized with ethanol and counterstained with DAPI or propidium iodide (PI). An experimental data set of several hundred double profiles was obtained by two parameter slit scan flow cytometry and evaluated automatically. The evaluation algorithm developed allowed a classification of chromosomes according to the number of centromeres and their chromosomal positions in less than 1 msec per individual profile. Approximately 20% of the measured DAPI profiles showed a bimodal distribution with a significant centromeric dip indicating a “ normal” chromosomal morphology and a correct alignment in the flow system. In many cases, profiles of a “normal” bimodal fluorescence distribution of the DNA stain (DAPI, PI) were correlated with a “ normal” FITC profile. Due to their centromeric indices these profiles agreed well to the expected human chromosomes of the cell line. In some cases of “ normal” DAPI (PI) profiles, “aberrant” FITC profiles were observed. These were interpreted as interspecies translocation chromosomes. For all results, there was a good agreement between flow cytometry and microscopic observations (digital image analysis)
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Juan, C., and J. Gosálvez. "Direct incorporation of fluorescein-12-dUTP to insect fixed chromosomes by random primed extension." Genome 37, no. 1 (February 1, 1994): 173–75. http://dx.doi.org/10.1139/g94-022.
Full textAbstract:
The use of an in situ random primed system allows the direct incorporation of fluorescein-12-dUTP into fixed insect chromosomes, resulting in a strong fluorescent labelling. While in an orthopteran species (Eyprepocnemis plorans) a relatively uniform labelling of meiotic and mitotic chromosomes is produced, in Tenebrio molitor (Coleoptera) only the chromosomal arms, but not the pericentromeric heterochromatic areas of mitotic chromosomes, show positive labelling by this method. This indicates that the organization of DNA in heterochromatin is distinct from that in the euchromatin in distantly related species, and in such a way that, in some species, the random hexanucleotides are prevented from annealing with the chromosomal denatured DNA or (and) that primer extension by Klenow enzyme is impeded.Key words: insect, chromosome, fluorescein-12-dUTP, in situ labelling.
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Runge, K. W., R. J. Wellinger, and V. A. Zakian. "Effects of excess centromeres and excess telomeres on chromosome loss rates." Molecular and Cellular Biology 11, no. 6 (June 1991): 2919–28. http://dx.doi.org/10.1128/mcb.11.6.2919.
Full textAbstract:
The linear chromosomes of eukaryotes contain specialized structures to ensure their faithful replication and segregation to daughter cells. Two of these structures, centromeres and telomeres, are limited, respectively, to one and two copies per chromosome. It is possible that the proteins that interact with centromere and telomere DNA sequences are present in limiting amounts and could be competed away from the chromosomal copies of these elements by additional copies introduced on plasmids. We have introduced excess centromeres and telomeres into Saccharomyces cerevisiae and quantitated their effects on the rates of loss of chromosome III and chromosome VII by fluctuation analysis. We show that (i) 600 new telomeres have no effect on chromosome loss; (ii) an average of 25 extra centromere DNA sequences increase the rate of chromosome III loss from 0.4 x 10(-4) events per cell division to 1.3 x 10(-3) events per cell division; (iii) centromere DNA (CEN) sequences on circular vectors destabilize chromosomes more effectively than do CEN sequences on 15-kb linear vectors, and transcribed CEN sequences have no effect on chromosome stability. We discuss the different effects of extra centromere and telomere DNA sequences on chromosome stability in terms of how the cell recognizes these two chromosomal structures.
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Runge, K. W., R. J. Wellinger, and V. A. Zakian. "Effects of excess centromeres and excess telomeres on chromosome loss rates." Molecular and Cellular Biology 11, no. 6 (June 1991): 2919–28. http://dx.doi.org/10.1128/mcb.11.6.2919-2928.1991.
Full textAbstract:
The linear chromosomes of eukaryotes contain specialized structures to ensure their faithful replication and segregation to daughter cells. Two of these structures, centromeres and telomeres, are limited, respectively, to one and two copies per chromosome. It is possible that the proteins that interact with centromere and telomere DNA sequences are present in limiting amounts and could be competed away from the chromosomal copies of these elements by additional copies introduced on plasmids. We have introduced excess centromeres and telomeres into Saccharomyces cerevisiae and quantitated their effects on the rates of loss of chromosome III and chromosome VII by fluctuation analysis. We show that (i) 600 new telomeres have no effect on chromosome loss; (ii) an average of 25 extra centromere DNA sequences increase the rate of chromosome III loss from 0.4 x 10(-4) events per cell division to 1.3 x 10(-3) events per cell division; (iii) centromere DNA (CEN) sequences on circular vectors destabilize chromosomes more effectively than do CEN sequences on 15-kb linear vectors, and transcribed CEN sequences have no effect on chromosome stability. We discuss the different effects of extra centromere and telomere DNA sequences on chromosome stability in terms of how the cell recognizes these two chromosomal structures.
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Heldrich, Jonna, Xiaoji Sun, Luis A. Vale-Silva, Tovah E. Markowitz, and Andreas Hochwagen. "Topoisomerases Modulate the Timing of Meiotic DNA Breakage and Chromosome Morphogenesis in Saccharomyces cerevisiae." Genetics 215, no. 1 (March 9, 2020): 59–73. http://dx.doi.org/10.1534/genetics.120.303060.
Full textAbstract:
During meiotic prophase, concurrent transcription, recombination, and chromosome synapsis place substantial topological strain on chromosomal DNA, but the role of topoisomerases in this context remains poorly defined. Here, we analyzed the roles of topoisomerases I and II (Top1 and Top2) during meiotic prophase in Saccharomyces cerevisiae. We show that both topoisomerases accumulate primarily in promoter-containing intergenic regions of actively transcribing genes, including many meiotic double-strand break (DSB) hotspots. Despite the comparable binding patterns, top1 and top2 mutations have different effects on meiotic recombination. TOP1 disruption delays DSB induction and shortens the window of DSB accumulation by an unknown mechanism. By contrast, temperature-sensitive top2-1 mutants exhibit a marked delay in meiotic chromosome remodeling and elevated DSB signals on synapsed chromosomes. The problems in chromosome remodeling were linked to altered Top2 binding patterns rather than a loss of Top2 catalytic activity, and stemmed from a defect in recruiting the chromosome remodeler Pch2/TRIP13 to synapsed chromosomes. No chromosomal defects were observed in the absence of TOP1. Our results imply independent roles for Top1 and Top2 in modulating meiotic chromosome structure and recombination.
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González-Siso, M. Isabel, M. Angeles Freire-Picos, M. Esperanza Cerdán, M. Wésolowski-Louvel, and H. Fukuhara. "Chromosomal mapping of the KlCYC1 gene from Kluyveromyces lactis." Genome 37, no. 3 (June 1, 1994): 515–17. http://dx.doi.org/10.1139/g94-073.
Full textAbstract:
Chromosomal assignment of the KlCYC1 gene from Kluyveromyces lactis has been performed by hybridization of the labelled probe to a DNA blot of isolated chromosomes. A clear hybridization signal to chromosome VI is reported.Key words: Kluyveromyces lactis, cytochrome c gene, chromosomal mapping.
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Vallente-Samonte, Rhea U., Robert A. Conte, and Ram S. Verma. "Syntenic homology of human unique DNA sequences within chromossome regions 5q31, 10q22, 13q32-33 and 19q13.1 in the great apes." Genetics and Molecular Biology 23, no. 3 (September 2000): 527–29. http://dx.doi.org/10.1590/s1415-47572000000300004.
Full textAbstract:
Homologies between chromosome banding patterns and DNA sequences in the great apes and humans suggest an apparent common origin for these two lineages. The availability of DNA probes for specific regions of human chromosomes (5q31, 10q22, 13q32-33 and 19q13.1) led us to cross-hybridize these to chimpanzee (Pan troglodytes, PTR), gorilla (Gorilla gorilla, GGO) and orangutan (Pongo pygmaeus, PPY) chromosomes in a search for equivalent regions in the great apes. Positive hybridization signals to the chromosome 5q31-specific DNA probe were observed at HSA 5q31, PTR 4q31, GGO 4q31 and PPY 4q31, while fluorescent signals using the chromosome 10q22-specific DNA probe were noted at HSA 10q22, PTR 8q22, GGO 8q22 and PPY 7q22. The chromosome arms showing hybridization signals to the Quint-EssentialTM 13-specific DNA probe were identified as HSA 13q32-33, PTR 14q32-33, GGO 14q32-33 and PPY 14q32-33, while those presenting hybridization signals to the chromosome 19q13.1-specific DNA probe were identified as HSA 19q13.1, PTR 20q13, GGO 20q13 and PPY 20q13. All four probes presumably hybridized to homologous chromosomal locations in the apes, which suggests a homology of certain unique DNA sequences among hominoid species.