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This research was the first report on karyological analysis and heteromorphic nucleolar organizer region of black surgeonfish (Acanthurus gahhm, Acanthuridae) in Thailand. The 10 male and 10 female specimens were collected from Phuket Marine Biological Center, and Phang Nga Coastal Research and Development Center, Andaman Sea, Thailand. Mitotic chromosomes were directly prepared from gill and kidney tissues. The chromosomes were stained by conventional Giemsa staining and Ag-NOR banding techniques. Results showed that the diploid chromosomes number of A. gahhm was 2n=48, the fundamental numbers (NF) was 54 in both male and female. The karyotype consist of 6 large acrocentric, 20 large telocentric, 18 medium telocentric and 4 small telocentric chromosomes. None of strange size chromosomes related to sex was found. The heteromorphic nucleolar organizer regions (NORs) were observed on telomeric short arm of first acrocentric which can defined as 1a1b. There is NOR in 1a and not in 1b. The karyotype formula of black surgeon fish was as follows: 2n (48) = La6+Lt20+Mt18+St4

The rDNA clusters and flanking sequences on human chromosomes 13, 14, 15, 21 and 22 represent large gaps in the current genomic assembly. The organization and the degree of divergence of the human rDNA units within an individual nucleolar organizer region (NOR) are only partially known. To address this lacuna, we previously applied transformation-associated recombination (TAR) cloning to isolate individual rDNA units from chromosome 21. That approach revealed an unexpectedly high level of heterogeneity in human rDNA, raising the possibility of corresponding variations in ribosome dynamics. We have now applied the same strategy to analyze an entire rDNA array end-to-end from a copy of chromosome 22. Sequencing of TAR isolates provided the entire NOR sequence, including proximal and distal junctions that may be involved in nucleolar function. Comparison of the newly sequenced rDNAs to reference sequence for chromosomes 22 and 21 revealed variants that are shared in human rDNA in individuals from different ethnic groups, many of them at high frequency. Analysis infers comparable intra- and inter-individual divergence of rDNA units on the same and different chromosomes, supporting the concerted evolution of rDNA units. The results provide a route to investigate further the role of rDNA variation in nucleolar formation and in the empirical associations of nucleoli with pathology.

The karyotype of the Odontocete whale, Mesoplodon densirostris, has not been previously reported. The chromosome number is determined to be 2n = 42, and the karyotype is presented using G-, C-, and nucleolar organizer region (NOR) banding. The findings include NOR regions on 2 chromosomes, regions of heterochromatic variation, a large block of heterochromatin on the X chromosome, and a relatively large Y chromosome. The karyotype is compared to published karyograms of 2 other species of Mesoplodon.

Chromosomal markers of the diversity and evolution of dinoflagellates are scarce because the genomes of these organisms are unique among eukaryotes in terms of their base composition and chromosomal structure. Similarly, a lack of appropriate tools has hindered studies of the chromosomal localization of 5S ribosomal DNA (rDNA) in the nucleosome-less chromosomes of dinoflagellates. In this study, we isolated and cloned 5S rDNA sequences from various toxin-producing species of the genus Alexandrium and developed a fluorescence in situ hybridization (FISH) probe that allows their chromosomal localization. Our results can be summarized as follows: 1) The 5S rDNA unit is composed of a highly conserved 122-bp coding region and an intergenic spacer (IGS), the length and sequence of which are variable even within strains. 2) Three different IGS types, one containing the U6 small nuclear RNA (snRNA) gene, were found among four of the studied species (A. minutum, A. tamarense, A. catenella and A. pacificum). 3) In all strains investigated by FISH (A. minutum, A. tamarense, A. pacificum, A. catenella, A. andersonii and A. ostenfeldii), 5S rDNA gene arrays were separate from the nucleolar organizer region, which contains the genes for the large 45S pre-ribosomal RNA. 4) One to three 5S rDNA sites per haploid genome were detected, depending on the strains/species. Intraspecific variability in the number of 5S rDNA sites was determined among strains of A. minutum and A. pacificum. 5) 5S rDNA is a useful chromosomal marker of mitosis progression and can be employed to differentiate vegetative (haploid) vs. planozygotes (diploid) cells. Thus, the FISH probe (oligo-Dino5Smix5) developed in this study facilitates analyses of the diversity, cell cycle and life stages of the genus Alexandrium., DIANAS, DETECCIÓN INNOVADORA DE PROLIFERACIONES ALGALES TÓXICAS: UNA NECESIDAD FRENTE AL CALENTAMIENTO GLOBAL, SI

This study investigated C-heterochromatin and NOR (nucleolar organizer region) characteristics of 2n = 52 and 2n = 60 chromosomal forms of Nannospalax xanthodon (blind mole rat), which are separate...

The genes of rRNA in the nucleolar organizer region (NOR) are inactivated in the oocytes of adult birds despite the functioning of lampbrush chromosomes. The nucleolus is not formed during all stages of the oocyte development. On the other hand, two morphological forms of oocytes differing by the presence of nucleolus in the germinal vesicle are described in the ovaries of juvenile birds. The activation and function of the ribosomal genes in avian oogenesis is still vague. In this work, the NOR activation in chicken (Gallus gallus domesticus) oocytes is confirmed with the help of fluorescence immunohistochemistry (antibodies against nucleophosmin, fibrillarin, and UBF1) and in situ nucleic acid hybridization (FISH with the probe to ITS1 in pre-rRNA). It is demonstrated that the nucleolus in the oocytes at the lampbrush stage in the chicken ovaries is fragmented after complete inactivation of the ribosome genes: the nucleolar fragments contain fibrillarin but do not contain pre-rRNA molecule. The utility of the ovary 3D reconstruction using serial histological sections for quantification of sex cell population heterogeneity in the ovaries of juvenile birds is demonstrated. The obtained results improve the current insight into the functional NOR state in the oocytes of juvenile female birds and contribute to the concept of diversity in the scenarios of gametogenesis.

This is chromosomal analysis of the brahminy blind snake, Ramphotyphlops braminus (Daudin, 1803), a newly recorded species in northeastern Thailand. Mitotic chromosomes were prepared directly from intestinal epithelia cells of five brahminy blind snakes. Conventional staining and Ag-NOR banding techniques were applied to stain the chromosomes. The results showed that the triploid chromosome number was 3n = 42, while the fundamental number (NF) was 78. The types of chromosomes were 24 macrochromosomes (19 m + 5sm) and 18 microchromosomes (9 m + 3sm + 6 t). The region adjacent to the short arm near the telomere of one in macrochromosome pair 8 showed a clearly observable nucleolar organizer region/NOR. There were no irregularly sized chromosomes related to sex. The karyotype formula for R. braminus is as follows: 3n (42) = Lm 12 + Sm 7 + Ssm 5 + 18 microchromosomes.

The objective of this study was to determine the variation in the number and size of nucleolar organizer regions (NOR) in the chinchilla karyotype. The study was performed with 12 standard chinchillas of two different lines. NORs were visualized on chromosome preparations by Ag-NOR silver staining. Four NOR size classes (I-IV) were determined on the basis of the results obtained, ranging from 0.070 (class I) to 0.229 (class IV). The mean NOR size was 0.144 µm² (SD=0.031 µm²) and fell within class II (from 0.101 to 0.150 µm²). Differences in the relative silver deposit area between the NOR-bearing pair of chromosomes were significant for 3 animals (P < 0.01) and for 1 animal (P < 0.05). The mean number of NORs in the animals ranged from 1.4 to 2.0 (SD=0.00–0.55). It was lower for chinchillas from central Poland (1.53±0.50) compared to those from southern Poland (1.68±0.48), with no significant differences (P > 0.05). The variation observed in the NOR size and number in the chinchilla karyotype indicates the occurrence of NOR polymorphism in the population.

The reference genome sequence of wheat 'Chinese Spring' (CS) is now available (IWGSC RefSeq v1.0), but the core sequences defining the nucleolar organizer regions (NORs) have not been characterized. We estimated that the total copy number of the rDNA units in the wheat genome is 11 160, of which 30.5%, 60.9% and 8.6% are located on Nor-B1 (1B), Nor-B2 (6B) and other NORs, respectively. The total length of the NORs is estimated to be 100 Mb, corresponding to approximately 10% of the unassembled portion of the genome not represented in RefSeq v1.0. Four subtypes (S1-S4) of the rDNA units were identified based on differences within the 3' external transcribed spacer regions in Nor-B1 and Nor-B2, and quantitative PCR indicated locus-specific variation in rDNA subtype contents. Expression analyses of rDNA subtypes revealed that S1 was predominantly expressed and S2 weakly expressed, in contrast to the relative abundance of rDNA subtypes in the wheat genome. These results suggest a regulation mechanism of differential rDNA expression based on sequence differences. S3 expression increased in the ditelosomic lines Dt1BL and Dt6BL, suggesting that S3 is subjected to chromosome-mediated silencing. Structural differences were detected in the regions surrounding the NOR among homoeologous chromosomes of groups 1 and 6. The adjacent regions distal to the major NORs were expanded compared with their homoeologous counterparts, and the gene density of these expanded regions was relatively low. We provide evidence that these regions are likely to be important for autoregulation of the associated major NORs as well as silencing of minor NORs.

Nucleoli are the sites of ribosome biogenesis and the largest membraneless subnuclear structures. They are intimately linked with growth and proliferation control and function as sensors of cellular stress. Nucleoli form around arrays of ribosomal gene (rDNA) repeats also called nucleolar organizer regions (NORs). In humans, NORs are located on the short arms of all five human acrocentric chromosomes. Multiple NORs contribute to the formation of large heterochromatin-surrounded nucleoli observed in most human cells. Here we will review recent findings about their genomic architecture. The dynamic nature of nucleoli began to be appreciated with the advent of photodynamic experiments using fluorescent protein fusions. We review more recent data on nucleoli in Xenopus germinal vesicles (GVs) which has revealed a liquid droplet-like behavior that facilitates nucleolar fusion. Further analysis in both XenopusGVs and Drosophila embryos indicates that the internal organization of nucleoli is generated by a combination of liquid-liquid phase separation and active processes involving rDNA. We will attempt to integrate these recent findings with the genomic architecture of human NORs to advance our understanding of how nucleoli form and respond to stress in human cells.

Hoplias malabaricus comprises seven karyomorphs (A-G) and evolutionary units have been described in some of them. In this study, the karyotypic composition and genomic organization of individual H. malabaricus from Central Amazon are described and to verify whether they can be classified according to known karyomorphs. Individuals from the Ducke Reserve have 2n = 42 chromosomes, similar to karyomorph A. Individuals from Catalão Lake and Marchantaria Island exhibit 2n = 40 chromosomes, similar to karyomorph C. Regarding the constitutive heterochromatin, individuals from all locations present centromeric/pericentromeric blocks, in addition to some bitelomeric and interstitial markings. The number of chromosomes with nucleolar organizer region, 5S rDNA and 18S rDNA sites varied among the different locations. The Rex 3 element has a compartmentalized distribution at the terminal and centromeric regions of most chromosomes, with subtle differences among populations. Fluorescence in situ hybridization performed with a telomeric probe allowed the detection of these regions only at the terminal ends of the chromosomes. Thus, only the chromosomal macrostructure (karyomorphs A-G) is not sufficient to establish evolutionary units within the H. malabaricus group, considering differences in the genome organization that are found among their populations. Such differences in the genomic organization could be mainly caused by the sedentary habits of this species.

The nucleolus is an important nuclear structure where transcription of ribosomal DNA (rDNA) takes place. During mitotic division, the nucleolus passes through different processes that inactivate rDNA transcription; in meiosis, its reassembly takes place during telophase II. The objective of this study was to identify the activity patterns and localization of nucleolar organizer regions (NORs) during meiotic division in fish species of the family Curimatidae. For this analysis, the meiotic division in five curimatid species was studied using silver nitrate impregnation, fluorescent in situ hybridization (FISH), and base-specific fluorochrome staining. Silver nitrate staining indicated the presence of a nucleolus in interphase nuclei, one chromosome pair in the spermatogonial metaphases, and one bivalent at the pachytene stage. No Ag-NORs were identified for cells at the diplotene, diakinesis, metaphase I, or metaphase II stages; however, FISH confirmed the presence of Ag-NORs in the nuclei, in spermatogonia, and at the pachytene phase. FISH identified this region during the other stages of meiosis, as did fluorochrome CMA3 staining, which revealed fluorescent marks corresponding to NORs during all stages of meiosis analyzed. The gene activity and localization of this ribosomal sequence during the different stages involved will also be discussed.

Trisomy mapping is a powerful method for assigning genes to chicken microchromosome 16 (GGA 16). The single chicken nucleolar organizer region (NOR), the 2 major histocompatibility complex regions (MHC-Y and MHC-B), and CD1 genes were all previously assigned to GGA 16 using trisomy mapping. Here, we combined array comparative genomic hybridization with trisomy mapping to screen unassigned genomic scaffolds (consigned temporarily to chrUn_random) for sequences originating from GGA 16. A number of scaffolds mapped to GGA 16. Among these were scaffolds that contain genes for olfactory (OR) and cysteine-rich domain scavenger (SRCR) receptors, along with a number of genes that encode putative immunoglobulin-like receptors and other molecules. We used high-resolution cytogenomic analyses to confirm assignment of OR and SRCR genes to GGA 16 and to pinpoint members of these gene families to the q-arm in partially overlapping regions between the centromere and the NOR. Southern blots revealed sequence polymorphism within the OR/SRCR region and linkage with the MHC-Y region, thereby providing evidence for conserved linkage between OR genes and the MHC within birds. This work localizes OR genes to the vicinity of the chicken MHC and assigns additional genes, including immune defense genes, to GGA 16.

Gossypium mustelinum ((AD)4) is one of five disomic species in Gossypium. Three 45S ribosomal DNA (rDNA) loci were detected in (AD)4 with 45S rDNA as probe, and three pairs of brighter signals were detected with genomic DNA (gDNA) of Gossypium D genome species as probes. The size and the location of these brighter signals were the same as those detected with 45S rDNA as probe, and were named GISH-NOR. One of them was super-major, which accounted for the fact that about one-half of its chromosome at metaphase was located at chromosome 3, and other two were minor and located at chromosomes 5 and 9, respectively. All GISH-NORs were located in A sub-genome chromosomes, separate from the other four allopolyploid cotton species. GISH-NOR were detected with D genome species as probe, but not A. The greatly abnormal sizes and sites of (AD)4 NORs or GISH-NORs indicate a possible mechanism for 45S rDNA diversification following (AD)4 speciation. Comparisons of GISH intensities and GISH-NOR production with gDNA probes between A and D genomes show that the better relationship of (AD)4 is with A genome. The shortest two chromosomes of A sub-genome of G. mustelinum were shorter than the longest chromosome of D sub-genome chromosomes. Therefore, the longest 13 chromosomes of tetraploid cotton being classified as A sub-genome, while the shorter 13 chromosomes being classified as D sub-genome in traditional cytogenetic and karyotype analyses may not be entirely correct. Wu Q, Liu F, Li S, Song G, Wang C, Zhang X, Wang Y, Stelly D, Wang K (2013) Uniqueness of the Gossypium mustelinum genome revealed by GISH and 45S rDNA FISH. J. Integr. Plant Biol. 55(7), 654–662.

Genetic variability was assessed among three species of genus Channa, namely C. gachua, C. punctata and C. striata, using karyological tools including karyomorphology, silver nitrate and chromomyci...

The work was aimed at determining the number and morphology of nucleoli in the prophase of the first meiotic division in domestic cattle males. The use of AgNO3 staining, commonly applied in cytogenetics for the identification of nucleolar organiser regions, made it possible to identify nucleoli in first-order spermatocytes. One nucleolus was identified in each analysed cell. Considerable morphological differentiation of the nucleoli during the prophase of the first meiotic division, particularly in leptotene, unobserved in other farm animal species, was noticed. Dark-hued grain-like structures were found within the disintegrating nucleoli, corresponding approximately or exactly to the number of the nucleolar organiser regions in the domestic cattle karyotype. Dark areas were identified in the selected prometaphase chromosomes. Their number corresponded with the number of active NORs defined in the domestic cattle karyotype.

Classical and molecular cytogenetic (18S rDNA, telomeric sequence, and LINE-1 retrotransposon probes) studies were carried out to contribute to an understanding of the organization of repeated DNA elements in the Amazon River dolphin (boto, Inia geoffrensis). Twenty-seven specimens were examined, each presenting 2n = 44 chromosomes, the karyotype formula 12m + 14sm + 6st + 10t + XX/XY, and fundamental number (FN) = 74. C-positive heterochromatin was observed in terminal and interstitial positions, with the occurrence of polymorphism. Interstitial telomeric sequences were not observed. The nucleolar organizer region (NOR) was located at a single site on a smallest autosomal pair. LINE-1 was preferentially distributed in the euchromatin regions, with the greatest accumulation on the X chromosome. Although the karyotype structure in cetaceans is considered to be conserved, the boto karyotype demonstrated significant variations in its formula, heterochromatin distribution, and the location of the NOR compared to other cetacean species. These results contribute to knowledge of the chromosome organization in boto and to a better understanding of karyoevolution in cetaceans.

The functional status of epithelial cells in inflammatory diseases of the cervix has been studied with the use of cytogenetic methods of detection of chromosome nucleolar organizers. The highest level of proliferation and rRNA synthesis was detected in cylindrical epithelial cells using indexes of compact and transitional nucleolonemic types of nuclear organizer regions; an elevated level of proliferation and rRNA synthesis was found in the squamous epithelial cells of the intermediate layer, and a low level of proliferation and rRNA synthesis was found in the squamous epithelial cells of the superficial layer.

Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:25:52Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:36:15Z : No. of bitstreams: 1 2-s2.0-79955837005.pdf: 1440943 bytes, checksum: d443947854f5d01dd9477e5845232d5c (MD5) Made available in DSpace on 2014-05-27T11:25:52Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-05-16 We made a cytogenetic analysis of four species of Oxyopidae and compared it with the karyotype data of all species of this family. In Hamataliwa sp, the mitotic cells showed 2n♂ = 26+X 1X 2 and telocentric chromosomes. The 2n♂ = 28, which has been described for only one oxyopid spider, is the highest diploid number reported for this family. Peucetia species exhibited distinct karyotype characteristics, i.e., 2n♂ = 20+X 1X 2 in P. flava and 2n♂ = 20+X in P. rubrolineata, revealing interspecific chromosome variability within this genus. However, both Peucetia species exhibited telocentric chromosomes. The most unexpected karyotype was encountered in Oxyopes salticus, which presented 2n♂ = 10+X in most individuals and a predominance of biarmed chromosomes. Additionally, one male of the sample of O. salticus was heterozygous for a centric fusion that originated the first chromosomal pair and exhibited one supernumerary chromosome in some cells. Testicular nuclei of Hamataliwa sp and O. salticus revealed NORs on autosomal pairs, after silver impregnation. The majority of Oxyopidae spiders have their karyotype differentiated by both reduction in diploid number chromosome number and change of the sex chromosome system to X type; however, certain species retain the ancestral chromosome constitution 2n = 26+X1X2. The most remarkable karyotype differentiation occurred in O. salticus studied here, which showed the lowest diploid number ever observed in Oxyopidae and the second lowest registered for Entelegynae spiders. © FUNPEC-RP www.funpecrp.com.br. Departamento de Biologia Universidade Estadual Paulista, Rio Claro, SP Departamento de Ciências Biológicas Universidade Federal de São Paulo, Diadema, SP Laboratório de Artrópodes, Instituto Butantan, São Paulo, SP Departamento de Biologia Universidade Estadual Paulista, Rio Claro, SP

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T19:05:05Z No. of bitstreams: 1 S1415-47572011000400006.pdf: 1383181 bytes, checksum: c699bc747ac131105daba4b851195d66 (MD5) Made available in DSpace on 2013-08-22T19:05:05Z (GMT). No. of bitstreams: 1 S1415-47572011000400006.pdf: 1383181 bytes, checksum: c699bc747ac131105daba4b851195d66 (MD5) Previous issue date: 2011-01-01 Made available in DSpace on 2013-09-30T20:08:48Z (GMT). No. of bitstreams: 2 S1415-47572011000400006.pdf: 1383181 bytes, checksum: c699bc747ac131105daba4b851195d66 (MD5) S1415-47572011000400006.pdf.txt: 28486 bytes, checksum: 6f63fe9b747905d7d6f5899d5372b96e (MD5) Previous issue date: 2011-01-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:51:35Z No. of bitstreams: 2 S1415-47572011000400006.pdf: 1383181 bytes, checksum: c699bc747ac131105daba4b851195d66 (MD5) S1415-47572011000400006.pdf.txt: 28486 bytes, checksum: 6f63fe9b747905d7d6f5899d5372b96e (MD5) Made available in DSpace on 2014-05-20T13:51:35Z (GMT). No. of bitstreams: 2 S1415-47572011000400006.pdf: 1383181 bytes, checksum: c699bc747ac131105daba4b851195d66 (MD5) S1415-47572011000400006.pdf.txt: 28486 bytes, checksum: 6f63fe9b747905d7d6f5899d5372b96e (MD5) Previous issue date: 2011-01-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) In this work, we analyzed the karyotypes of five species. Hypostomus cf. heraldoi, from the Mogi-Guaçu River, had 2n = 72 chromosomes, with a nucleolar organizer region (NOR) in one chromosomal pair. Hypostomus regani, from the Mogi-Guaçu River had 2n = 72 chromosomes with NORs in two chromosomal pairs. Hypostomus sp., from the Mogi-Guaçu River basin, had 2n = 68 chromosomes, with NORs in two chromosomal pairs. Hypostomus aff. agna, from Cavalo Stream, had 2n = 74 chromosomes with NORs in two chromosomal pairs. Hypostomus cf. topavae, from Carrapato Stream, had 2n = 80 chromosomes, with NORs in two chromosomal pairs. Hypostomus species showed marked diversity in the karyotypic formula, which suggested the occurrence of several Robertsonian rearrangements and pericentric inversions during the evolutionary history of this genus. This hypothesis was supported by the occurrence of a large number of uniarmed chromosomes and multiple NORs in a terminal position in most species and may be a derived condition in the Loricariidae. Universidade Estadual Paulista Instituto de Biociências Departamento de Morfologia Universidade Estadual de Maringá (UEM) Departamento de Biologia Universidade Estadual Paulista Instituto de Biociências Departamento de Morfologia CNPq: 140644/2005-9 CNPq: 300314/2003-5 CNPq: 306066/2009-2

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T19:05:04Z No. of bitstreams: 1 S1415-47572010000400012.pdf: 265909 bytes, checksum: ece273c362162eb8a4f54377d1e17b9a (MD5) Made available in DSpace on 2013-08-22T19:05:04Z (GMT). No. of bitstreams: 1 S1415-47572010000400012.pdf: 265909 bytes, checksum: ece273c362162eb8a4f54377d1e17b9a (MD5) Previous issue date: 2010-01-01 Made available in DSpace on 2013-09-30T20:08:45Z (GMT). No. of bitstreams: 2 S1415-47572010000400012.pdf: 265909 bytes, checksum: ece273c362162eb8a4f54377d1e17b9a (MD5) S1415-47572010000400012.pdf.txt: 21574 bytes, checksum: 986cbda8d638201be35fcf2939ad6cb7 (MD5) Previous issue date: 2010-01-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:54:49Z No. of bitstreams: 2 S1415-47572010000400012.pdf: 265909 bytes, checksum: ece273c362162eb8a4f54377d1e17b9a (MD5) S1415-47572010000400012.pdf.txt: 21574 bytes, checksum: 986cbda8d638201be35fcf2939ad6cb7 (MD5) Made available in DSpace on 2014-05-20T13:54:49Z (GMT). No. of bitstreams: 2 S1415-47572010000400012.pdf: 265909 bytes, checksum: ece273c362162eb8a4f54377d1e17b9a (MD5) S1415-47572010000400012.pdf.txt: 21574 bytes, checksum: 986cbda8d638201be35fcf2939ad6cb7 (MD5) Previous issue date: 2010-01-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Theridiidae is a derived family within the Araneoidea clade. In contrast to closely related groups, the 2n(male) = 20+X1X2 with acro/telocentric chromosomes is the most widespread karyotype among the theridiid spiders. In this work, the cytogenetic analysis of Argyrodes elevatus revealed original chromosome features different from those previously registered for Theridiidae, including the presence of 2n(male) = 20+X with meta/submetacentric chromosomes. Most individuals of Nesticodes rufipes showed family conserved karyotype characteristics. However, one individual had a 2n(male) = 24 due to the presence of an extra chromosome pair, which exhibited regular behavior and reductional segregation during meiosis. After silver staining, mitotic cells exhibited NORs localized on the terminal regions of the short arms of pairs 2, 3, and 4 of A. elevatus and on the terminal regions of long arms of pair 4 of N. rufipes. The comparative analysis with data from phylogenetically related species allowed the clarification of the origin of the interspecific and intraspecific chromosome variability observed in Argyrodes and in N. rufipes, respectively. Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Biologia Universidade Federal de São Paulo (UNIFESP) Departamento de Ciências Biológicas Universidade Estadual de Mato Grosso do Su Instituto Butantan Laboratório de Artrópodes Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Biologia FAPESP: 06/53275-3 FAPESP: 08/55633-0

Meiotic behavior, pollen morphology, interphasic nucleus pattern and karyotype description for Senna occidentalis (Caesalpinioideae — Fabaceae) are presented. The species had non-reticulate interphasic nucleus and homogeneous chromosome condensing with minute distal late-condensing portions in prometaphase. In mitotic metaphase, chromosome number was 2n = 28 (9 m + 5 sm), with secondary constriction in one chromosome pair. Detection of four nucleoli indicated existence of two chromosome pairs bearing Nucleolar Organizer Region (NOR). Meiotic behavior was regular, with high meiotic index (95%). Pollen grains, classified as polar/spheroidal, presented 90% viability.

We examined chromosomal distribution of major ribosomal DNAs (rDNAs), clustered in the nucleolar organizer regions (NORs), in 18 species of moths and butterflies using fluorescence in situ hybridization (FISH) with a codling moth (Cydia pomonella) 18S rDNA probe. Most species showed one or two rDNA clusters in their haploid karyotype but exceptions with four to eleven clusters also occurred. Our results in a compilation with previous data revealed dynamic evolution of rDNA distribution in Lepidoptera except Noctuoidea, which showed a highly uniform rDNA pattern. In karyotypes with one NOR, interstitial location of rDNA prevailed, whereas two-NOR karyotypes showed mostly terminally located rDNA clusters. A possible origin of the single interstitial NOR by fusion between two NOR-chromosomes with terminal rDNA clusters lacks support in available data. In some species, spreading of rDNA to new, mostly terminal chromosome regions was found. The multiplication of rDNA clusters without alteration of chromosome numbers rules out chromosome fissions as a major mechanism of rDNA expansion. Based on rDNA dynamics in Lepidoptera and considering the role of ordered nuclear architecture in karyotype evolution, we propose ectopic recombination, i.e. homologous recombination between repetitive sequences of non-homologous chromosomes, as a primary motive force in rDNA repatterning.

The karyotype of the teleostean Gymnothorax tile was analysed by replication banding in order to clearly identify the homologous chromosomes. The RBG pattern was then compared to those of the other Muraenidae studied (G. unicolor and Muraena helena) and banding homologies of nine chromosome pairs were pointed out. A single nucleolar organizer region (NOR) was localized by FISH, silver- and CMA3-staining on the short arm of the pair 12. Telomeric (TTAGGG) repeats were terminally localized by FISH on all the chromosome pairs. The genome size and the AT-DNA content were evaluated by flow cytometry. Available cytogenetic data on the Muraenidae were then compared and discussed.

Analyses of nucleolar organizer region (NOR) site polymorphism using silver staining were carried out in 72 individuals of 5 geographically isolated populations of Tor putitora from India. All Ag-NORs analyzed were constituted by rDNA that was confirmed by the presence of positive fluorescence in situ hybridization (FISH) signals. Analyses of the extra NORs among the populations confirmed the hypothesis that multi-chromosomal NOR site polymorphism is not an exception but a rule. We found a negative correlation between the mean number of extra NORs and geographical distance from the Alaknanda River population, and a significant positive correlation between the genetic distances obtained by NOR data and geographical distances among the isolated populations. These findings suggest a single geographical origin of this NOR polymorphism, from which it might have radiated to neighboring populations before their isolation. The distribution of NOR site variation was compared with the sequence variation of the ITS1 region of major rDNA (45S) and the two were found to be positively related. Different mechanisms such as transposable elements adjacent to ribosomal genes, repetitive elements which could serve as a point for chromosome exchange, amplification of minor rDNA loci, and reinsertion of extrachromosomal rDNA amplified during oogenesis have been proposed to explain the apparent NOR site polymorphism. The high dispersion of NOR site variants within and among the populations suggests that whatever transposition phenomenon is responsible for repetitive DNA or NOR jumping, it could still be active in T. putitora.

In the present study, we examined the chromosome number and morphology, heterochromatin pattern, and location of the nucleolar organizer region (NOR) and telomeric sequences in Aplastodiscus albofrenatus, A. arildae, A. ehrhardti and A. eugenioi in order to cytogenetically characterize the Aplastodiscus albofrenatus group. The 4 species analyzed in this study had diploid numbers of 2n = 22 and very similar chromosome morphology. These species could be differentiated based on the distribution and amount of heterochromatin and the location of the nucleolar organizer region (NOR). Six of the 8 A. albofrenatus individuals had an NOR polymorphism previously unknown in anurans since only one of the homologs of pairs 1 and 7 was stained. In the other 2 specimens, the NOR occurred on both homologs of pair 7. In A. ehrhardti, pairs 6 and 10 were stained by the AgNOR technique, but only pair 6 was confirmed by in situ hybridization. The NOR was located on pair 10 in A. arildae and on pair 7 in A. eugenioi. In A. albofrenatus and A. arildae, multivalent rings involving NOR-containing chromosomes were observed during prophase I of meiosis. The telomeric probe identified the telomeres in all species and also centromeric regions in the chromosomes of A. albofrenatus and A. arildae, which were coincident with centromeric heterochromatin. The conserved chromosomal morphology seen mainly in the first 7 pairs among species of the A. albofrenatus group suggests that all of these species probably originated from a common ancestral karyotype and that chromosomal rearrangements resulted in karyotype differentiation, with changes in NOR location, as well as telomeric and heterochromatin dispersal.

Pseudis paradoxa paradoxa, P. p. platensis, P. bolbodactyla, P. fusca and P. tocantins were analyzed cytogenetically by conventional chromosomal staining, C-banding, silver staining and fluorescent in situ hybridization with an rDNA probe. Pseudis tocantins chromosomes were also stained with distamycin A/DAPI. All of the species had a diploid number of 2n = 24 chromosomes and the nucleolar organizer region (NOR) was located on pair 7. However, the karyotypes could be differentiated based on the morphology of chromosomal pairs 2 and 8, the region that the NORs occupied on the long arms of the homologous of pair 7, and the pattern of heterochromatin distribution. The subspecies P. p. paradoxa and P. p. platensis had identical karyotypes. Heteromorphism in NOR size was seen in P. p. paradoxa, P. p. platensis, P. bolbodactyla and P. fusca. Heteromorphic sex chromosomes (ZZ/ZW) were identified in P. tocantins. The W chromosome was subtelocentric and larger than the metacentric Z chromosomes. The differences observed in the C-banding pattern and in the position of the NOR on the sex chromosomes suggested that inversions and heterochromatinization were responsible for the morphological differentiation of these chromosomes.

Aim: The synthesis of rRNA is a key determinant of normal and malignant cell growth and subject to epigenetic regulation. Yet, the epigenomic features of rDNA arrays clustered in nucleolar organizer regions are largely unknown. We set out to explore for the first time how DNA methylation is distributed on individual rDNA arrays. Materials & Methods: Here we combined immunofluorescence detection of DNA modifications with fluorescence hybridization of single DNA fibers, metaphase immuno-FISH and methylation-sensitive restriction enzyme digestions followed by Southern blot. Results: We found clustering of both hypomethylated and hypermethylated repeat units and hypermethylation of noncanonical rDNA in IMR90 fibroblasts and HCT116 colorectal carcinoma cells. Surprisingly, we also found transitions between hypo- and hypermethylated rDNA repeat clusters on single DNA fibers. Conclusion: Collectively, our analyses revealed co-existence of different epialleles on individual nucleolar organizer regions and showed that epi-combing is a valuable approach to analyze epigenomic patterns of repetitive DNA.

Nucleoli are the product of the activity of nucleolar organizer regions (NOR) in certain chromosomes. Their main functions are the formation of ribosomal subunits from ribosomal protein molecules and the transcription of genes encoding rRNA. The aim of the study was to determine the shape of nucleoli and analyse methylation in the gene RN28S in the spermatocytes of male Japanese quail (Coturnixjaponica) in two age groups. Nucleoli were analysed in cells of the first meiotic prophase. Their number and shape were determined and they were classified as regular, irregular or defragmented. In the cells of the young birds no defragmented nucleoli were observed, with regular and irregular nucleoli accounting for 97% and 3%, respectively. In the cells of older birds no regular nucleoli were observed, while irregular and defragmented nucleoli accounted for 37% and 67%, respectively. MSP (methylation-specific PCR) showed that the gene RN28S is methylated in both 15-week-old and 52-week-old quails. In recent years an association has been established between nucleolus morphology and cellular ageing processes.

The electric eel, Electrophorus electricus, the only species of its genus, has a wide distribution in the Amazon and Orinoco drainages. There is little previous information regarding the population variation in E. electricus, with only basic karyotype data from two populations (Amazon and Araguaia Rivers). Karyotypic description and analysis of CO1 barcode sequences were performed for E. electricus from three localities (Caripetuba, Irituia, and Maicuru Rivers). All samples share the 2n=52 (42 m-sm [meta-submetacentric] +10 st-a [subtelo-acrocentric]) with previously studied material. However, the Maicuru River samples differ from the other populations, as they have B chromosomes. The distribution of noncentromeric constitutive heterochromatin between samples is relatively divergent. All samples analyzed present the Nucleolar Organizer Region (NOR) located in a single chromosome pair. In the samples from Caripetuba, NORs were colocalized with a heterochromatin block, whereas the NOR was flanked by heterochromatin in Maicuru River samples and pericentromeric heterochromatin adjacent NOR was found in Irituia River samples. Alignment of CO1 barcode sequences indicated no significant differentiation between the samples analyzed. Results suggest that karyotypic differences between samples from the Caripetuba, Irituia, and Amazon Rivers represent chromosome polymorphisms. However, differences between the samples from the Maicuru and Araguaia Rivers and the remaining populations could represent interpopulation differentiation, which has not had time to accrue divergence at the CO1 gene level.

Different cytogenetic techniques including C-banding, base-specific fluorochromes and silver nitrate staining were used to compare the karyotypes of three species of bugs, representatives of the Neotropical genus Antiteuchus, namely A. mixtus, A. sepulcralis and A. macraspis. The species have holokinetic chromosomes which is typical of the Hemiptera, and their diploid chromosome number is 2n = 14 and they have an XX/XY (female/male) sex chromosome system. C-banding revealed telomeric blocks of constitutive heterochromatin in most chromosomes of these three species. In each species, CMA;-positive blocks were observed in the telomeric heterochromatin region of chromosome pair 1. This chromosome pair carried the nucleolar organizer region (NOR), which silver nitrate staining revealed is located in the telomeric heterochromatin. In contrast to the result obtained with CMA3, entire chromosome complement stained homogenously with DAPI, except for the telomeric region of pair 1, which remained unstained. Fil: Lanzone, Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto Argentino de Investigaciones de las Zonas Áridas. Provincia de Mendoza. Instituto Argentino de Investigaciones de las Zonas Áridas. Universidad Nacional de Cuyo. Instituto Argentino de Investigaciones de las Zonas Áridas; Argentina. Universidade Federal de Pernambuco; Brasil Fil: de Souza, Maria José. Universidade Federal de Pernambuco; Brasil

Cytogenetic studies were carried out on samples of Parapteronotus hasemani, Sternarchogiton preto and Sternarchorhamphus muelleri (Apteronotidae, Gymnotiformes) from the Amazon basin. The first two species exhibited both a 2n = 52 karyotype, but differed in their karyotypic formulae, distribution of constitutive heterochromatin, and chromosomal location of the NOR. The third species, Sternarchorhamphus muelleri, was found to have a 2n = 32 karyotype. In all three species the DAPI and chromomycin A3 staining results were consistent with the C-banding results and nucleolar organizer region (NOR) localization. The 18S rDNA probe confirmed that there was only one pair of ribosomal DNA cistron bearers per species. The telomeric probe did not reveal interstitial telomeric sequences (ITS). The karyotypic differences among these species can be used for taxonomic identification. These data will be useful in future studies of these fishes and help understanding the phylogenetic relationships and chromosomal evolution of the Apteronotidae.

Meiotic chromosomes obtained from members of the coleopteran subfamilies Rutelinae and Dynastinae were studied using standard and silver nitrate staining, C-banding, base-specific fluorochromes and fluorescent in situ hybridization (FISH). The study presents detailed karyotipic descripitions of three Rutelinae species (Geniates borelli, Macraspis festiva and Pelidnota pallidipennis), and two Dynastinae species (Lygirus ebenus and Strategus surinamensis hirtus) with special emphasis on the distribution and variability of constitutive heterochromatin and the nucleolar organizer region (NOR). We found that for G. borelli, P. pallidipennis, L. ebenus and S. s hirtus the karyotype was 2n = 20 (9II + Xy p), with G. borelli, P. pallidipennis and L. ebenus showed meta-submetacentric chromosomes which gradually decreased in size. For Macraspis festiva the karyotype was 2n = 18 (8II + Xy p). In L. ebenus we found that the NOR was located on an autosome, but in the other four species it occurred on the sex bivalents. In all five species the constitutive heterochromatin (CH) was predominantly pericentromeric while the X chromosomes were almost completely heterochomatic, although CMA3/DA/DAPI staining showed intra and interspecific variation in the bright fluorescence of the constitutive heterochromatin. The FISH technique showed rDNA sites on the X chromosome of the Rutelinae species.

We used silver nitrate staining, CMA3 and FISH to study the chromosomal localization of both the major ribosomal genes and the nucleolar organizer regions as well as that of the minor ribosomal genes (5S rDNA) in two species of Batoidea, Taeniura lymma (Dasyatidae) and Raja montagui (Rajidae). In both species, all the metaphases examined showed the presence of multiple NOR-bearing sites, while the gene for 5S rRNA proved to be localized on two chromosome pairs. Furthermore, one of the two 5S rDNA sites in T. lymma was shown to be co-localized with the major ribosomal cluster. The presence of multiple nucleolar organizer regions in the two species might be interpreted as being the result of intraspecific polymorphisms, or as a phenomenon of the amplified transposition of mobile elements of the genome. We also determined the nucleotide sequence of the 5S rRNA gene, consisting of 564 bp in R. montagui and 612 bp in T. lymma. We also found TATA-like and (TGC) n trinucleotides, (CA) n dinucleotides and (GTGA) n tetranucleotides, which probably influence gene regulation.

Six loci of nucleolar organizer region (NOR) were detected in genomicin situ hybridization (GISH) of cotton (Gossypium). NOR was the characteristic of 45S rDNA but could be generated by genomic DNA (gDNA) extracted fromGossypium species as probe. With twice FISH to the same mitotic cell ofG. herbaceum orG. hirsutum, number, position and size for NORs generated from 45S rDNA and gDNA were identified largely similar or even the same. The NORs with gDNA as probe were therefore permanently defined as GISH-NORs. GISH-NORs fromG. hirsutum andG. raimondii mitotic images were all terminal types. Four and two GISH-NORs fromG. herbaceum (var.africanum) were terminal and centromere types, respectively. Six GISH-NORs inG. hirsutum were chromosome mapped with two in A- and four in D-subgenomes. There were also GISH-NORs in mitotic image ofG. raimondii with its own gDNA as probe. From mitotic image ofG. herbaceum with its own gDNA as probe, GISH-NOR could not be observed but non-whole-recovery of hybridized signals was distinguished. These non-whole-recovery of hybridized signals were detected on long arm terminals of most chromosomes and especially existed in nearly half long arm of a pair of chromosomes inG. herbaceum gDNA probed itself GISH image, which may be possibly induced by low copy genes within the regions rather than inter-subgenomic segment translocations. GISH-NORs in G.hirsutum mitotic images were dominantly observed when gDNAs from D and A genome species were used as probes and block, respectively, but not when the reverse probe and block gDNA from the two diploid progenitor genomes were designed. There may be two speculations to this special phenomenon: rDNA concerted evolution; content of rDNA in genome D more than genome A.

Mitotic cells from three Brassica species, B. campestris L., B. oleracea L., and B. napus L. have been differentially stained after cold treatment and application of the ASG method for G-banding. Karyotypes and idiograms are presented. In all three species the chromosomes can be matched to individual pairs. The heterochromatic satellite next to the nucleolar organizer region (NOR) differs from the rest of the chromosomal material by having a very faint colour. It responds to the staining procedure in the same way as cold sensitive regions of chromatin described in other species. In Brassica this type of reaction is apparently limited to the satellite next to the active NOR.

The number and loci of nucleolar organizer regions (NOR) on chromosomes in Sika deer (Cervus nippon centralis) were determined by fluorescence in situ hybridization with a human 28S ribosomal RNA (rRNA) gene as a probe. Sika deer that live in Nikko National Park and its neighboring areas (Asio and Seta) in Japan were used. All of the analyzed metaphases had three or four NOR at the end of the first and second longest telocentric autosomes. Nucleolar organizer region association, which is associated specifically on parts of NOR between chromosomes, was also observed clearly. A Sika deer 28S rRNA gene was produced by a polymerase chain reaction method. The nucleotide sequence of a Sika deer 28S rRNA gene determined by an automatic sequencer was 97 bp, and showed homogeneity of 88% for the human sequence.

The purpose of this work was to study the karyotype, spermatogenesis and nucleolar activity at meiosis, in the species Rhodnius domesticus (Heteroptera, Triatominae). The testicular tubules were cytologically prepared by the conventional method of cell crushing and subsequent application of cytogenetic staining techniques with lacto-acetic orcein and silver-ion impregnation. The species under study presented karyotype 2n = 20A+XY, the modal number of the subfamily Triatominae. The chromosomes presented no primary constriction and were therefore characterized as holocentric. It was observed that the sex chromosomes sometimes were located at the periphery, close to the ring formed by autosomes, at first meiotic division. At metaphases II, sex chromosomes were positioned in the center of the autosomal ring, thus evidencing a postreductional behavior. These same chromosomes showed late migration at anaphases and were clearly impregnated with silver-ions, suggesting they bore Nucleolar Organizer Reg...