Differences in the rDNA-bearing chromosome divide the Asian-Pacific and Atlantic species of Crassostrea (Bivalvia, Mollusca)

Karyotype and chromosomal location of the major ribosomal RNA genes (rDNA) were studied using fluorescence in situ hybridization (FISH) in five species of CRASSOSTREA: three Asian-Pacific species (C. gigas, C. plicatula, and C. ariakensis) and two Atlantic species (C. virginica and C. rhizophorae). FISH probes were made by PCR amplification of the intergenic transcribed spacer between the 18S and 5.8S rRNA genes, and labeled with digoxigenin-11-dUTP. All five species had a haploid number of 10 chromosomes. The Atlantic species had 1-2 submetacentric chromosomes, while the three Pacific species had none. FISH with metaphase chromosomes detected a single telomeric locus for rDNA in all five species without any variation. In all three Pacific species, rDNA was located on the long arm of Chromosome 10 (10q)--the smallest chromosome. In the two Atlantic species, rDNA was located on the short arm of Chromosome 2 (2p)--the second longest chromosome. A review of other studies reveals the same distribution of NOR sites (putative rDNA loci) in three other species: on 10q in C. sikamea and C. angulata from the Pacific Ocean and on 2p in C. gasar from the western Atlantic. All data support the conclusion that differences in size and shape of the rDNA-bearing chromosome represent a major divide between Asian-Pacific and Atlantic species of CRASSOSTREA: This finding suggests that chromosomal divergence can occur under seemingly conserved karyotypes and may play a role in reproductive isolation and speciation.     

Cytogenetic analysis in Polypterus ornatipinnis (Actinopterygii, Cladistia, Polypteridae) and 5S rDNA

Polypteridae is a family of archaic freshwater African fish that constitute an interesting subject for the study of the karyological evolution in vertebrates, on account of their primitive morphological characters and peculiar relationships with lower Osteichthyans. In this paper, a cytogenetic analysis on twenty specimens of both sexes of Polypterus ornatipinnis the ornate "bichir", coming from the Congo River basin, was performed by using both classical and molecular techniques. The karyotypic formula (2n=36; FN=72) was composed of 26 M+10 SM. The Alu I banding, performed to characterize heterochromatin in this species, was mainly centromeric. Both the chromosome location of the ribosomal 5S and 18S rRNA genes were examined by using Ag-NOR, classical C-banding, CMA(3) staining and FISH. CMA(3) marked all centromerical regions and showed the presence of two GC rich regions on the p arm of the chromosome pair n°1 and on the q arm of the pair n°14. Staining with Ag-NOR marked the only telomeric region of the chromosome n°1 p arm. After PCR, the 5S rDNA in this species was cloned, sequenced and analyzed. In the 665bp 5S rDNA sequence of P.ornatipinnis, a conserved 120bp gene region for the 5S rDNA was identified, followed by a non-transcribed variable spacer (NTS) which included simple repeats, microsatellites and a fragment of a non-LTR retrotransposon R-TEX. FISH with 5S rDNA marked the subtelomeric region of the q arm of the chromosome pair n°14, previously marked by CMA(3). FISH with 18S rDNA marked the telomeric region of the p arm of the pair n°1, previously marked both by Ag-NOR and CMA(3). The (GATA)(7) repeats marked the telomeric regions of all chromosome pairs, with the exclusion of the n°1, n°3 and n°14; hybridization with telomeric probes (TTAGGG)(n) showed signals at the end of all chromosomes. Karyotype evolution in Polypterus genus was finally discussed, including the new data obtained.     

Cytogenetic and molecular characterization of the Abies alba genome and its relationship with other members of the Pinaceae

Genome size, karyotype structure, heterochromatin distribution, position and number of ribosomal genes, as well as the ITS2 sequence of the internal transcribed spacer (ITS) were analysed in silver fir (Abies alba Mill.). The analysis also included characterization of the Arabidopsis-type of telomeric repeats in silver fir and in related species. The results were compared with results from other species of the Pinaceae, to evaluate phylogeny and chromosomal and molecular evolution in the Pinaceae. Integrated chromosomal data provided insights into chromosome and karyotype evolution in the Pinaceae. The evolutionary trend for GC-rich heterochromatic blocks seems to involve loss of blocks that are not associated with rDNA. Similarly, numerous large blocks of interstitial plant telomeric repeats that are typical for all analysed species of the genus Pinus were not observed in the evolutionarily younger genera, such as Abies, Picea and Larix. On the contrary, the majority of telomeric sequences in these three genera appeared confined to the chromosome ends. We confirmed the current position of Abies and Tsuga in subfamily Abietoideae and the position of Pinus in the subfamily Pinoideae based on ITS2 sequences. Pseudotsuga is placed together with Larix into the subfamily Laricoideae. We conclude that the current position of the genus Picea in the subfamily Abietoideae should be reconsidered and, possibly, the genus Picea should be reclassified as a separate subfamily, Piceoideae, as recently proposed.     

Methylated-rich regions and tandem repeat arrays along the chromosome complement of Colpodium versicolor (Stev.) Schmalh

The grass Colpodium versicolor (Stev.) Schmalh is one of six angiosperms with extremely reduced chromosome set 2n?=?2x?=?4. The chromosome complement of this species was studied. The DNA methylation pattern was determined with a specific monoclonal antiboby. 5-Methylcytosine residues are present in different chromosomal sites, with specific occurrence, some methylated bands showing differences between homologous chromosomes. Moreover, a fluorescent in situ hybridisation with telomere repeats and 45S rDNA sequences were performed. Hybridisation signals of telomeric repeats are detectable at the distal ends of the two pair of chromosomes, while 45S rDNA is localised in one chromosomal site, corresponding to the secondary constriction. In addition, 45S rDNA, as well as telomere-associated sequences, results to be 5-methylcytosine-enriched. The results are discussed and compared with those previously obtained in other plant systems 2n?=?4 with the aim to enable a better knowledge of the lengthwise differentiation of this chromosome complement.     

Telomeric DNA allocation in chromosomes of common shrew (Sorex araneus, eulipotyphla)

Recently, we displayed an Iberian shrew species (Sorex granarius) with telomere structures unusual for mammals. Long telomeres on the short acrocentric arms contain an average of 213 kb of telomere repeats, whereas the other chromosomal ends have only 3.8 kb (Zhdanova et al., 2005; 2007). However, it is not clear whether these telomeres are typical for all shrew species or only for S. granarius. S. granarius and common shrew Sorex araneus are sibling species. In this study, using modified Q-FISH we demonstrated that telomeres in S. araneus from various chromosomal races distinguished by their number of metacentrics contain 6.8–15.2 kb of telomeric tracts. The S. araneus telomere lengths appear to correspond to telomere lengths in the majority of both shrew species and wild mammals, whereas S. granarius has telomeres with unique or rare structures. Using DNA and RNA high-specific modified probes to telomeric repeats (PNA and LNA), we showed that interstitial telomeric sites in S. araneus chromosomes contain mainly telomeric DNA and that their localization coincide with some evolutionary breakpoints. Interstitial telomeric DNA in S. granarius chromosomes was not revealed. Thus, the distribution of telomeric DNA may be significantly different, even in closely related species whose chromosomes are composed of almost identical chromosomal arms.     

Intrachromosomal location of the telomeric repeat (TTAGGG)n

Eukaryotic telomeres are specialized DNA-protein structures that are thought to ensure chromosomal stability and complete replication of the chromosome ends. All telomeres which have been studied consist of a tandem array of G-rich repeats which seem to be sufficient for telomere function. Originally, the human telomeric repeat (TTAGGG)_n was assumed to be exclusively located at the very end of all human chromosomes. More recent evidence, however, suggests an extension into proterminal regions. Very little is known about the interstitial distribution of telomeric repeats. Here we present evidence for the presence of (TTAGGG)_n repeats in internal loci on the long and short arms of different human chromosomes. In addition, we studied the genomic organization of these repeats in more detail and discuss possible functions of interstitial telomeric repeats in the human genome.     

Ribosomal, telomeric and heterochromatin sequences localization in the karyotype of Anemone hortensis

The karyotype of the Mediterranean species Anemone hortensis L. (Ranunculaceae) was characterized with emphasis on heterochromatin distribution and localization of ribosomal (18S−5.8S−26S and 5S rDNA) and telomeric repeats (TTTAGGG). Diploid chromosome complement, 2 n  = 2 x  = 16, common to all investigated populations, consisted of three acrocentric, one meta-submetacentric and four metacentric chromosomes ranging in size from 6.34 to 10.47 µm. Fluorescence in situ hybridization (FISH) with 18S and 5S rDNA probes revealed two 18S−5.8S−26S rDNA loci on a satellite and secondary constriction of acrocentric chromosome pair 2 and terminally on acrocentric chromosome pair 3, and two 5S rDNA loci in the pericentromeric region of meta-submetacentric chromosome pair 4 and in the proximity of the 18S−5.8S−26S rDNA locus on chromosome pair 2. The only GC-rich heterochromatin, as revealed by fluorochrome Chromomycin A3 staining, was that associated with nucleolar organizer regions, whereas AT-rich heterochromatin, stained with 4,6-diamino-2-phenylindole (DAPI), was distributed intercalarly and terminally on the long arm of all three acrocentric chromosomes, and terminally on chromosomes 4 and 5. FISH with Arabidopsis -type telomeric repeats (TTTAGGG) as a probe revealed two classes of signals, small dot-like and large bands, at chromosome termini exclusively, where they corresponded to terminal DAPI-stained heterochromatin. Heteromorphism of chromosome pair 4, which refers to terminal DAPI bands and FISH signals, was observed in populations of Anemone hortensis . Chromosome pairing during meiosis was regular with formation of localized chiasmata proximal to the centromere.  © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society , 2006, 150 , 177–186.     

Constitutive heterochromatin, 5S and 18S rDNA genes in Apareiodon sp. (Characiformes, Parodontidae) with a ZZ/ZW sex chromosome system

Karyotype, sex chromosome system and cytogenetics characteristics of an unidentified species of the genus Apareiodon originating from Piquiri River (Paraná State, Brazil) were investigated using differential staining techniques (C-banding and Ag-staining) and fluorescent in situ hybridization (FISH) with 5S and 18S rDNA probes. The diploid chromosome number was 2n = 54 with 25 pairs of meta- (m) to submetacentric (sm) and 2 pairs of subtelocentric (st) chromosomes. The major ribosomal rDNA sites as revealed by Ag-staining and FISH with 18S rDNA probe were found in distal region of longer arm of st chromosome pair 26, while minor 5S sites were observed in the interstitial sites on chromosome pairs 2 (smaller cluster) and 7 (larger one). The C-positive heterochromatin had pericentromeric and telomeric distribution. The heteromorphic sex chromosome system consisted of male ZZ (pair 21) and female middle-sized m/st Z/W chromosomes. The pericentric inversion of heterochromatinized short arm of ancestral Z followed by multiplication of heterochromatin segments is hypothesized for origin of W chromosome. The observed karyotype and chromosomal markers corresponded to those found in other species of the genus.     

Sequence comparison of distal and proximal ribosomal DNA arrays in rice (Oryza sativa L.) chromosome 9S and analysis of their flanking regions

Rice (Oryza sativa ssp. japonica cv. Nipponbare) harbors a ribosomal RNA gene (rDNA) cluster in the nucleolar-organizing region at the telomeric end of the short arm of chromosome 9. We isolated and sequenced two genomic clones carrying rice rDNA fragments from this region. The rice rDNA repeat units could be classified into three types based on length, which ranged from 7,928 to 8,934 bp. This variation was due to polymorphism in the number of 254-bp subrepeats in the intergenic spacer (IGS). Polymerase chain reaction (PCR) analysis suggested that the rDNA units in rice vary widely in length and that the copy number of the subrepeats in the IGS ranges from 1 to 12 in the rice genome. PCR and Southern blot analyses showed that most rDNA units have three intact and one truncated copies of the subrepeats in the IGS, and distal (telomere-side) rDNA units have more subrepeats than do proximal (centromere-side) ones. Both genomic clones we studied contained rDNA-flanking DNA sequences of either telomeric repeats (5′-TTTAGGG-3′) or a chromosome-specific region, suggesting that they were derived from the distal or proximal end, respectively, of the rDNA cluster. A similarity search indicated that retrotransposons appeared more frequently in a 500-kb portion of the proximal rDNA-flanking region than in other subtelomeric regions or sequenced regions of the genome. This study reveals the repetitive nature of the telomeric end of the short arm of chromosome 9, which consists of telomeric repeats, an rDNA array, and a retrotransposon-rich chromosomal region.Sequence accession numbers in DDBJ assigned for OSJNOa063K24 and OSJNBb0013K10 are AP009051 and AP008245, respectively.     

Do time, heterochromatin, NORs, or chromosomal rearrangements correlate with distribution of interstitial telomeric repeats in Sigmodon (cotton rats)?

We studied the chromosomal distribution of telomere repeats (TTAGGG)(n) in 8 species of Sigmodon (cotton rats) using chromosome paints fluorescent in situ hybridization (FISH) from Sigmodon hispidus. In 2 species with the proposed primitive karyotype for the genus, telomere repeats were restricted to telomeric sites. But in the other 6 species that include 3 with proposed primitive karyotypes and 3 with highly rearranged karyotypes, telomere repeats were found on both telomeric sites and within interstitial telomeric sites (ITSs). To explain the distribution of ITS in Sigmodon, we gather data from C-bands, silver nitrate staining, G-bands, and chromosomal paint data from previous published studies. We did find some correlation with ITS and heterochromatin, euchromatic chromosomal rearrangements, and nucleolar organizing regions. No one type of chromosomal structure explains all ITS in Sigmodon. Multiple explanations and mechanisms for movement of intragenomic sequences are required to explain ITS in this genus. We rejected the hypothesis that age of a lineage correlates with the presence of ITS using divergence time estimate analyses. This multigene phylogeny places species with ITS (S. arizonae, S. fulviventer, S. hispidus, S. mascotensis, S. ochrognathus, and S. toltecus) in the clade with a species without ITS (S. hirsutus). Lineages with ITS (S. arizonae and S. mascotensis) arose independently from a lineage absent of ITS (S. hirsutus) around 0.67 to 0.83 Ma. The rearranged karyotypes of S. mascotensis and S. arizonae appear to be an independently derived autapomorphic characters, supporting a fast rate of chromosomal changes that vary among species.     

Comparative cytogenetics in four species of Palinuridae: B chromosomes, ribosomal genes and telomeric sequences

The evolutionary pathway of Palinuridae (Crustacea, Decapoda) is still controversial, uncertain and unexplored, expecially from a karyological point of view. Here we describe the South African spiny lobster Jasus lalandii karyotype: n and 2n values, heterochromatin distribution, nucleolar organizer region (NOR) location and telomeric repeat structure and location. To compare the genomic and chromosomal organization in Palinuridae we located NORs in Panulirus regius, Palinurus gilchristi and Palinurus mauritanicus: all species showed multiple NORs. In J. lalandii NORs were located on three chromosome pairs, with interindividual polymorphism. In P. regius and in the two Palinurus species NORs were located on two chromosome pairs. In the two last species 45S ribosomal gene loci were also found on B chromosomes. In addition, the nature and location of telomeric repeats were investigated by FISH in J. lalandii, P. gilchristi, P. mauritanicus Palinurus elephas, and P. regius (Palinuridae, Achelata), and in Scyllarus arctus (Scyllaridae, Achelata): all these Achelata species showed the (TTAGG)n pentameric repeats. Furthermore, in J. lalandii these repeats occurred in all the telomeres and in some interstitial chromosomal sites, associated with NORs.     

Fluorescence in situ hybridization of rDNA, telomeric (TTAGGG)n and (GATA)n repeats in the red abalone Haliotis rufescens (Archaeogastropoda: Haliotidae)

The physical location of 18S-5.8S-28S rDNA, telomeric sequences with (TTAGGG)n DNA probe and (GATA)n microsatellites were performed by fluorescence in situ hybridization in chromosomes of red abalone Haliotis rufescens. The karyotype of red abalone showed a diploid number of 36 (8M+9SM+1ST). FISH performed with rDNA probe, showed the location of major ribosomal clusters in the terminal region of the large arms of two submetacentric pairs (chromosome 4 and 5). Localization of heteromorphisms of FISH-rDNA was found between chromosome homologues and sister chromatids in all metaphases analyzed. This indicates that rDNA clusters are variable within the red abalone genome. The variability in the NOR-bearing reported using silver staining in other gastropods and our result are discussed. In addition, the presence of microsatellite (TTAGGG)n and (GATA)n was demonstrated after FISH treatment by DNA probes. The telomeric sequence occurred at the ends of all mitotic chromosomes, while the (GATA)n repetitive was found on chromosomal interstitial zones as well as at the telomeres in abalone chromosomes.     

3D organization of interphase fibroblast nuclei in two closely related shrew species, Sorex granarius and S. araneus, differed by the structure of chromosome termini

To study 3D organization of fibroblast interphase nuclei in two sibling shrew species, Sorex araneus from Cordon race and S. granarius, FISH with probe to telomeric and rDNA repeats, and immunofluorescence with ANA CREST and antibodies to nucleolus protein B23 were used. Karyotypes of studied species are composed of near identical chromosomal arms and differ by the number of metacentrics and the structure of terminal chromosome regions. The large telomeres containing on the average 218 kbp of telomere repeats characterize the short arms in all of 32 S. granarius acrocentrics. Telomere repeats in them alternate with nbosomal repeats. These regions also contain active NORs. In contrast, active NORs in S. araneus are localized at the terminal regions of 8 chromosomal arms (Zhdanova et al., 2005, 2007b). We have shown that telomere associations of chromosomes and contacts of a part of telomere clusters with inner nuclear membrane and nucleolus characterize interphase nuclei of both S. granarius and S. araneus. Moreover, the partial colocalization of telomere and ribosomal clusters, and spatial nearness of centomeric and telomeric regions were revealed in the interphase nuclei of S. granarius. Evidently, only those ribosomal clusters that contain a number of active ribosomal genes display connection with nucleolus. The stripping of nucleolus materials during transition of fibroblasts to mitosis and the role of B23 protein in this process has been studied.     

Mapping of the 18S and 5S ribosomal RNA genes in the fish Prochilodus argenteus Agassiz, 1829 (Characiformes,Prochilodontidae)

A single NOR-bearing chromosome pair was identified by silver nitrate staining in a previous study of the fish Prochilodus argenteus from the S ã o Francisco River (MG, Brazil), with a third metacentric chromosome sporadically bearing active NOR. The present study focused on an analysis of the chromosomal localization of both the major (45S) and the minor (5S) rRNA genes using FISH. The use of the 18S rDNA probe confirmed the previous Ag-NOR sites interstitially located in a large metacentric pair and also identified up to three other sites located in the telomeric regions of distinct chromosomes, characterizing an interindividual variation of these sites. In addition, the 5S rDNA site was revealed adjacent to the major NOR site, identified at the end of the large Ag-NOR bearing metacentric chromosome. In a few metaphases, an additional weak hybridization signal was observed in a third chromosome, possibly indicating the presence of another 5S rDNA cluster. Despite a lower karyotype diversification (2n=54 and FN=108) often observed among species of Prochilodontidae, variations involving both 45S and 5S rRNA genes could play an important role in their chromosome diversification.     

Comparative cytogenetic analysis of the “Sylvaemus” group of Apodemus (Rodentia, Muridae): A. sylvaticus from Sicily and A. flavicollis from the central Apennines

Chromosome mapping of three common markers, ie major and 5S rRNA genes (rDNA) and telomeric repeats, and conventional chromosome bandings were applied to two sibling species, Apodemus sylvaticus Linnaeus, 1758 and A. flavicollis Melchior, 1834, to further investigate intra- and interspecific karyological differentiation in the genus Apodemus. A slight variation of the rDNA-patterns was detected between the two Apodemus species. In both of them, the major NORs were located on autosome pairs 8, 11, 12 and 22, while the two other rDNA sites detected on chromosomes 7 and 21 were variable in, respectively, A. sylvaticus and A. flavicollis. Several tiny rDNA sites were present on the sex chromosomes in both species, but their incidence was lower in A. flavicollis. Single 5S rDNA chromosomal sites were conserved on chromosome pair 20. No interstitial sites of telomeric repeats were present in either species. In the Sicilian population of A. sylvaticus, the constitutive heterochromatin pattern corresponded to the “sylvaticus-E1” cytotype, while A. flavicollis had a species-specific pattern restricted to centromeres of all chromosomes. The results are discussed in relation to cytogenetic data available for the genus, with emphasis on the Sylvaemus group/subgenus.     

C-Banding/DAPI and in situ hybridization reflect karyotype structure and sex chromosome differentiation in Humulus japonicus Siebold & Zucc

Japanese hop (Humulus japonicus Siebold & Zucc.) was karyotyped by chromosome measurements, fluorescence in situ hybridization with rDNA and telomeric probes, and C-banding/DAPI. The karyotype of this species consists of sex chromosomes (XX in female and XY1Y2 in male plants) and 14 autosomes difficult to distinguish by morphology. The chromosome complement also shows a rather monotonous terminal distribution of telomeric repeats, with the exception of a pair of autosomes possessing an additional cluster of telomeric sequences located within the shorter arm. Using C-banding/DAPI staining and 5S and 45S rDNA probes we constructed a fluorescent karyotype that can be used to distinguish all autosome pairs of this species except for the 2 largest autosome pairs, lacking rDNA signals and having similar size and DAPI-banding patterns. Sex chromosomes of H. japonicus display a unique banding pattern and different DAPI fluorescence intensity. The X chromosome possesses only one brightly stained AT-rich terminal segment, the Y1 has 2 such segments, and the Y2 is completely devoid of DAPI signal. After C-banding/DAPI, both Y chromosomes can be easily distinguished from the rest of the chromosome complement by the increased fluorescence of their arms. We discuss the utility of these methods for studying karyotype and sex chromosome evolution in hops.     

Localization of rDNA sites in holocentric chromosomes of three species of triatomines (Heteroptera, Triatominae)

Chromatin organization in the holocentric chromosomes of three triatomines species was cytologically studied by fluorescent in situ hybridization with a 45S rDNA probe of Drosophila melanogaster to localize ribosomal genes. In Triatoma tibiamaculata, metaphases I showed telomeric highlights in a single, larger bivalent. In T. protacta, hybridization was detected in one of the telomeres of an autosomal chromosome. In T. platensis, there were highlights in a single, smaller chromosome (X chromosome). The results obtained did not agree with the expected localization of rDNA genes in the sex chromosomes of triatomines, as demonstrated by silver impregnation, and suggest that the chromosome reorganization that occurred in this group during evolution may be a more important mechanism involved in rDNA distribution.     

Cytogenetic characterization of the sole Solea senegalensis (Teleostei: Pleuronectiformes: Soleidae): Ag-NOR, (GATA) n , (TTAGGG) n and ribosomal genes by one-color and two-color FISH

A cytogenetic analysis of the sole Solea senegalensis was carried out using silver staining for the nucleolus organizer region (Ag-NOR) identification, one-color FISH for chromosomal mapping of 45S and 5S ribosomal DNAs (rDNAs), (GATA) _n , and (TTAGGG) _n , and two-color FISH for co-localization of both rDNAs. The Ag-NORs and the 45S rDNA were mapped to a medium-sized submetacentric chromosomal pair. Hybridization with the 5S rDNA showed a major signal on the short arm of a medium-sized submetacentric chromosome pair and a minor signal on a centromeric site of a small acrocentric chromosome pair. Differences in the Ag-NOR and 45S and 5S rDNAs FISH signal sizes were observed between homologous chromosomes and among individuals. A two-color FISH co-localized 45S and 5S rDNAs to a medium-sized submetacentric chromosomal pair. The hybridization with the telomeric (TTAGGG) _n repeat displayed small signals at all chromosomal telomeres. Finally, the (GATA) _n probe produced dispersed and small hybridization signals on all chromosome spreads, showing its ubiquitous existence in the genome. These results were compared with those from other Pleuronectiformes and discussed in terms of karyotype evolution.     

Karyotypic characterization and genomic organization of the 5S rDNA in <Emphasis Type="Italic">Polypterus senegalus</Emphasis> (Osteichthyes,Polypteridae)

Polypteridae (Cladistia) is a family of archaic fishes, confined to African freshwaters. On account of their primitiveness in anatomical and morphological characters and mosaic relationships among lower Osteichthyans fishes, they constitute an important subject for the study of evolution in vertebrates. Very little is known about the karyological structure of these species. In this article, a cytogenetic analysis on twenty specimens of Polypterus senegalus (Cuvier, 1829) was performed using both classical and molecular techniques. Karyotype (2n = 36; FN = 72), chromosome location of telomeric sequences (TTAGGG) _n , (GATA)₇ repeats and ribosomal 5S and 18S rRNA genes were examined by using Ag-NOR, classical C-banding, CMA₃ staining and FISH. Staining with Ag-NOR showed the presence of two GC rich NORs on the p arm of the chromosome pair no. 1. CMA₃ marked all centromerical and some (no. 1 and no. 14) telomeric regions. FISH with 5S rDNA marked the subtelomeric region of the q arm of the chromosome pair no. 14. FISH with 18S rDNA marked the telomeric region of the p arm of the chromosome pair no. 1, previously marked by Ag-NOR. (GATA)₇ repeats marked the subtelomeric regions of all chromosome pairs, with the exclusion of the no. 1, 3 and 14. Hybridization with telomeric probes (TTAGGG) _n showed bright signals at the end of all chromosomes. After cloning, the 5SrDNA alignment revealed an organization of sequences made up of two different classes of tandem arrays (5S type I and 5S type II) of different lengths.     

Chromosomal diversity in three species of electric fish (Apteronotidae,Gymnotiformes) from the Amazon Basin

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.