NOR Banding Research Articles

Evolutionary History of Procyonids (Carnivora: Procyonidae): Comparative Cytogenetics of Nasua Nasua and Procyon Cancrivorus

The chromosome number of all species of Procyonidae was very conserved. Most have very strong chromosomal homology among members of the family, as well as with different species of the suborder Feliformia. Many chromosomes have similar and sometimes identical G-band patterns, while others differ due to Robertsonian translocations and inversions. In this work, the karyotype of two species of the family Procyonidae, Nasua nasua (2n=38; NF = 74) and Procyon cancrivorus (2n=38; NF = 76), the latter described cytogenetically for the first time, is characterized using conventional staining and differential G, C and NOR banding techniques. Pericentric and paracentric inversions were observed in Procyon cancrivorus and Nasua nasua , respectively. Finally, the G-band karyotypes of both species were compared with the Ancestral Carnivore Karyotype proposed in the literature, providing a homology and conservation on the karyotype evolution of this order.

Comparative cytogenetic analysis of fishes in the genus Trichopodus (Osphronemidae) in Thailand

Abstract. Supiwong W, Wongchantra P, Thongnetr W, Mingkwan B, Chaiyasan P, Pinmongkhonkul S, Pinthong K, Tanomtong A. 2021. Comparative cytogenetic analysis of fishes in the genus Trichopodus (Osphronemidae) in Thailand. Biodiversitas 22: 3029-3036. Comparative cytogenetic study of four species of the genus Trichopodus including T. leerii, T. microlepis, T. pectoralis, and T. trichopterus from Thailand, was carried out. The specimens were collected from the Basins throughout Thailand. Chromosome preparation was directly performed from the kidney tissues. Conventional staining by Giemsa solution, Ag–NOR banding by silver nitrate solution and C-banding by NaOH solution were conducted to stain the chromosomes. Results showed that four species studied have the same diploid number and fundamental number as 46 composing of all telocentric chromosomes. Karyotypes consisting of large-medium-small sizes in T. leerii, T. microlepis, T. pectoralis and T. trichopterus were 16–28–2, 14–32–0, 20–26–0 and 10–30–6 chromosomes, respectively. The marker chromosomes which present the NOR positions are the single pair in all species but there are differences in the locations and pairs such as pairs no. 1, 7, 2 and 1, respectively. Most species except T. trichopterus, NOR sites locate at interstitial region adjacent to the centromere. T. trichopterus had the telomeric NORs. Constitutive heterochromatin blocks displayed at the centromeric/pericentromeric regions of all chromosomes in T. leerii and T. microlepis whereas in T. pectoralis and T. trichopterus, those presented at not only centromeres of several chromosome pairs, but they also were found at interstitial sites. The obtained finding, cytogenetic data has species-specific. Thus, it can be used for further study on taxonomy. Moreover, this data shows the close relationship of genetics among these species so that the application for breeding may be used in the future.

Classical and molecular cytogenetics of Belontia hasselti (Perciformes: Osphronemidae): Insights into the ZZ/ZW sex chromosome system

Abstract. Chaiyasan P, Mingkwan B, Jantarat S, Suwannapoom C, Cioffi MDB, Liehr T, Talumphai S, Tanomtong A, Supiwong W. 2021. Classical and molecular cytogenetics of Belontia hasselti (Perciformes: Osphronemidae): Insights into the ZZ/ZW sex chromosome system. Biodiversitas 22: 546-554. Karyotype of Java combtail fish, Belontia hasselti, from To Daeng peat swamp forest, Narathiwat Province, southern Thailand, was studied for the first time. Mitotic chromosome preparations were prepared directly from kidney cells from ten male and ten female fish. Conventional staining, NOR banding, and molecular cytogenetics with fluorescence in situ hybridization (FISH) using 5S and 18S rDNAs, as well as microsatellites d(CA)15 and d(CAC)10 as probes were applied. The diploid chromosome number (2n) was 48 and a female heterogametic sex chromosome system (ZZ/ZW) is suggested. The fundamental numbers (NF) were 48 and 49 in males and females, respectively. The karyotype of males comprised 48 telocentric chromosomes while the female ones were composed of one metacentric and 47 telocentric chromosomes. A single Ag-NOR-bearing chromosomal pair was identified. The NOR positions were characterized at the interstitial sub-centromeric region of pair 13, which coincided with signals of 18S rDNA and d(CAC)10 probes. The 5S rDNA signals were located at interstitial sites of the largest telocentric pair. Microsatellite d(CA)15 repeats were highly distributed throughout almost all entire chromosomes except for centromeric regions on some chromosome pairs, including sex chromosomes. The present study is a novel report for a ZZ/ZW sex chromosome system of this fish family in Thailand.

A Study of Constitutive Heterochromatin and NOR Banding in Three Species of Puntius from the State of Haryana, India

The purpose of the present study was to investigate and compare the karyotypes of three species of Puntius viz. Puntius sarana (Hamilton, 1822), Puntius sophore (Hamilton, 1822) and Puntius ticto (Hamilton, 1822) belonging to family Cyprinidae in terms of chromosomal architecture, banding pattern and number of chromosomes from aquatic ecosystems of Haryana, India. Diploid chromosome no. 50 was observed in all 3 species of Puntius. The chromosomes of 3 species of Puntius showed constitutive heterochromatin at telomeric and centromeric regions of chromosomes.The Ag-NOR (Argyrophilic-Nucleolus Organizer Region) bands were observed on homologous chromosome pair nos. 2, 8 and 14 in P. sophore, pair nos. 2, 9 and 14 in P. ticto. The diploid chromosome number in P. sarana was found to be 2n = 50, 2 pairs of metacentric chromosomes, 3 pairs of subtelocentric and 20 pairs of acrocentric chromosomes. Chromosomal studies on the P. sophore revealed the diploid chromosome number to be 2n = 50, showed 16 pairs of metacentric chromosomes, 5 pairs of submetacentric, 2 pairs of subtelocentric and 2 pairs of acrocentric chromosomes. The diploid chromosome number in P. ticto was found to be 2n = 50, with 15 pairs of metacentric chromosomes, 6 pairs of submetacentric, 2 pairs of subtelocentric and 2 pairs of acrocentric chromosomes. No heteromorphic sex chromosomes were cytologically detected. Variations in chromosomes are observed with respect to earlier studies which may be due to variation in habitat conditions as a result of anthropogenic activities.

Chromosomal Metaphase Spreads with G and NOR bandings of Fresh Water Teleost, Clarias batrachus

Background,The detail structure of chromosomes depicted by their banding pattern is very useful in drawing homology and analogy in the structure of chromosome within and between the species. Results, In the present study, the diploid number of chromosomes was 50 in the fresh water teleost, Clarias batrachus in which 18 metacentric, 20 submetacentric and 12 telocentric chromosomes were observed. Structurally, the G bands revealed that transverse bands were visible after banding. These were due to the presence of heterochromatin and presence and absence of bases, these bands were observed at two pairs of submetacentric chromosomes. The NOR bands on the chromosomes were detected by silver staining in the two pairs of medium size telocentric chromosomes. It was believed that silver staining bound with the non-histone proteins of the transcriptionally active NOR region. Conclusion, Our results showed that basic information of chromosome number and morphology with G and NOR banding can be used as an important tool for cytogenetic data of fresh water teleost Clarias batrachus.

Additional description of karyotype and meiotic features of Takydromus sexlineatus (Squamata, Lacertidae) from northeastern Thailand

This study determined the standardized karyotype and idiogram of Takydromus sexlineatus from Surin Province in northeastern Thailand. Mitotic metaphase chromosomes from bone marrow cells and different meiotic stage from testicular cells respectively were studied after staining with conventional Giemsa and Ag-NOR banding techniques for chromosome identification. The study was carried out with samples taken from both male and female T. sexlineatus, the sex of which was identified by external examination. The results showed that the diploid chromosome number of T. sexlineatus was 2n = 38 and the fundamental number (NF) was 38 in both male and female. Although chromosome study was done in both sexes of T. sexlineatus, this study does not identify the sex chromosomes. This is because of the limitation of the staining techniques used. Therefore, we did not conclude on the sex chromosome system in this species and all chromosomes were taken as autosomes. The types of autosomes observed were 16 large telocentric, 14 medium telocentric, and 6 small telocentric macrochromosomes and 2 microchromosomes. Our result is the new record in Thailand on this species and NOR characteristics by silver staining is the first report on NOR banding in this species. The karyotype formula of T. sexlineatus thus was found as: 2n(38) = L 16 t + M 14 t + S 6 t + 2 microchromosomes.

A study of constitutive heterochromatin and NOR banding in three species of Indian major carps from the State of Haryana, India

Cytogenetic survey of fishes becomes increasingly important to establish chromosomal relation between the teleosts, to have a glimpse of the relation between chromosomal evolution and differentiation of vertebrate species. In the present study, Chromosome banding studies was done in three species of carps i.e. Catla catla (Hamilton, 1822), Labeo rohita (Hamilton, 1822) and Cirrhinus mrigala (Hamilton, 1822). Diploid chromosome number 50 was observed in all 3 species of carps. The chromosomes of C. catla, L. rohita and C. mrigala showed constitutive heterochromatin at telomeric and centromeric regions of chromosomes. The Ag-NOR (Argyrophilic-Nucleolus Organizer Region) bands were observed on homologous chromosome pair number 11 in C. catla, 15th chromosome pair in L. rohita. In C. mrigala, the Ag- NOR staining elucidated the presence of darkly stained NORs on the terminal region of the long arms of one of the chromosome. Another homologue of this chromosome pair could not be localized due to scattering of chromosomes. The results depict that variation in ecological conditions with time due to human activities can not only affect the chromosomal number but also chromosomal morphology.

Cytogenetic and molecular studies in conservation breeding of Pulawska breed pigs

The paper presents the genetic characteristics of Pulawska breed pigs carried out on the basis of cytogenetic and molecular studies. Among chromosome markers, polymorphic variants of specific chromosome structures such as centromeric heterochromatin areas (C bands) and nucleolar organizing regions (NOR bands) were taken into account. The described breed-specific tendencies concerning size polymorphism of these structures are the source of chromosome markers that are useful for identifying the linkage with genes controlling important production traits. On the other hand, molecular studies presented in this work included DNA markers of the STR and SNP type (with particular consideration of the PRL, FST, MC4R, TNNT3, MTTP and DIO3 genes), which are useful in determining the genetic background of functional traits, as well as the characteristics of the breeds for terms of genetic variation, especially conservative breeds, where it is appropriate to maintain the existing genetic diversity and intra-breed variability.

Cytogenetic data on the threatened leafcutter ant Atta robusta Borgmeier, 1939 (Formicidae: Myrmicinae: Attini)

The karyotype of the threatened ant species Atta robusta is described so as to establish the evolutionary relationships of this taxon with other leafcutter ants. Standard Giemsa staining, C-banding, NOR banding, fluorochromes CMA3/DAPI, Hsc-FA technique and Fluorescence in situ Hybridization (FISH) using 18S rDNA probe were conducted on a population from Aracruz, state of Espírito Santo, Brazil, allowing for comparisons with data available on Atta and other fungus-growing ant species. The diploid chromosome number observed for A. robusta was 2n=22, and the karyotypic formula was 18m+2sm+2st. Heterochromatic blocks were observed in the centromeric region of most chromosomes, where one pair of metacentric chromosomes is characterized by a GC-rich heterochromatic band in the interstitial region of its long arm. The detection of 18S rDNA using FISH confirmed the presence of single NOR for A. robusta. This is the first report of rDNA 18S detection using FISH for leafcutter ants. The cytogenetic results of this study confirm the information available for Atta and allow us to confirm the conserved chromosome number, morphology and banding pattern within the genus for the taxa studied to date, which included species from three out of the four groups of Atta indicated by molecular data. The accumulation of cytogenetic data on fungus-growing ants enhances the understanding of the genomic evolutionary patterns of Atta, since it belongs to a group of recent origin between the most well studied ants. Cytogenetic data does not indicate restrictions in relocation or reintroduction in areas where populations were extinct due to the conserved karyotype. This study allows for cytogenetic comparison of A. robusta with other ants of Atta, emphasizing the importance of chromosomal information for species conservation.

Distribution of constitutive heterochromatin and nucleolar organizer regions in two species of family Gomphidae (Odonata: Anisoptera)

Spermatogonial and primary spermatocyte chromosomes of Paragomphus lineatus (Selys) and Nepogomphus modestus (Selys) of family Gomphidae have been described on the basis of C-banding and NOR staining. Both the species possess 2n (♂) =23, n (♂) =12 without m chromosomes with XO(♂)/XX(♀) type sex determining mechanism. The sex chromosome is the largest element in the whole complement in Paragomphus lineatus but it is medium sized in Nepogomphus modestus. In Paragomphus lineatus, all the autosomal bivalents possess terminal C-bands while X chromosome is C-positive throughout the length but lightly stained in the centre. Similarly, terminal NOR bands are present on all the autosomal bivalents and X chromosome shows terminal NOR bands with large interstitial band. In Nepogomphus modestus, all the autosomal bivalents except one bivalent possess terminal C-bands while X chromosome represents large C-positive region on one end and small C-negative region on the other end. NOR staining shows terminal / non terminal NOR bands on some autosomal bivalents while X chromosome possesses dark NOR band only on one end.

Investigation of Partamona helleri (Apidae, Meliponini) B chromosome origin. An approach by microdissection and whole chromosome painting

The stingless bee Partamona helleri in southeast Brazil shows the regular chromosome number 2n = 34 and a variable number of up to four minute B1 or B2 chromosomes. Previous cytogenetic analyses have indicated morphological similarities between the B1 chromosome and chromosome segments in the regular karyotype. In this study, microdissection and chromosome painting were employed along with C banding, NOR banding, and base-specific fluorochrome staining to investigate the origin of the B1 chromosome in P. helleri. B1-generated probe hybridized exclusively to B1 chromosomes. This result suggests an independent origin from the regular karyotype or, alternatively, that the B chromosome may have suffered substantial genetic alterations along its independent evolution. The absence of higher dosages of these small B chromosomes in this population of P. helleri may be related to the existence of either a genetic or cytogenetic constraint in the establishment of such high numbered karyotypes.

Banding and FISH in three species of Vernonia, subsection Macrocephalae (Asteraceae, Vernonieae)

To complement our knowledge about the karyotypes of the genus Vernonia Schreb., different techniques of chromosome banding, including AgNOR, triple staining with fluorochromes CMA/DA/DAPI (CDD), and fluorescence in situ hybridization (FISH) for the 45S rDNA probe, were applied to three species of subsection Macrocephalae. Vernonia bardanoides was collected from an area of cerrado (savanna) vegetation in Itirapina, Sao Paulo State, Brazil, and V. linearifolia and V. tomentella were collected from areas of rocky, open altitudinal vegetation in Joaquim Felicio and Diamantina, respectively, in Minas Gerais State. All species showed two terminal CMA+ and NOR bands. FISH indicated two terminal 45S rDNA sites in V. linearifolia and V. tomentella, and six in V. bardanoides.

45, X Karyotipe Sahip İnfertil Erkek Hastada Y;21 Translokasyonu

A 30-year-old male patient referred to our clinic for unraveling the underlying etiology of the azoospermia. He had no unusual medical history. At physical examination, obesity, short neck and gynecomastia were noted. All hormone levels were normal except estradiol which was 2-fold higher than the upper limit. Having azoospermia in the spermiogram, scrotal ultrasonography was normal. Cytogenetic analysis and fluorescence in situ hybridization were performed subsequently, 45,X,add(21)(p10) and 45,X,add(21)(p10).ish der(Y;21) (q12;p10) were found, respectively. On C-banding, dicentric staining of translocated chromosome was observed. NOR banding was negative. Molecular genetics studies using multiplex polymerase chain reaction revealed the presence of Y chromosome sequences at SRY, AZFa, AZFb, AZFc regions.

Karyotype characterization of two populations of Vernonia geminata (Asteraceae, Vernonieae) using banding and FISH techniques

In order to extend our knowledge concerning karyotypes of the genus Vernonia, we applied various techniques of chromosome banding, including AgNOR and triple staining with the fluorochromes CMA/DA/DAPI (CDD), and of fluorescent in situ hybridization (FISH) for the 45S rDNA probe to specimens of two populations of Vernonia geminata collected from an open-pasture area, in southern Brazil. B chromosomes were observed in one of the populations. Both populations of V. geminata presented a pair of CMA(3)(+) terminal bands and one pair of chromosomes with terminal AgNOR banding. The FISH evidenced, in one population, two pairs of small sites of 45S rDNA; these being two small terminal sites and two centromeric sites. In the other population, there was only one pair of small terminal sites and two sites in two B chromosomes, one in each chromosome. There was coincidence of localization between CMA(+) and NOR bands with one of the small terminal sites of 45S rDNA of one chromosome of the normal complement, but not in B chromosomes.

Karyotype and NOR-banding of mitotic chromosomes of some Vitis L. species

Chromosome studies were performed in V. champinii, V. cinerea, V. girdiana, V. labrusca, V. rotundifolia, V. rupestris and V. vinifera with the purpose of species characterization using chromosome morphometric data and NOR banding. A median ideogram was obtained for each species. The karyotype formula obtained varied from 7m + 12sm to 9m + 11sm. The species showed moderate chromosome asymmetry values according to TF% form, Stebbins, Romero Zarco and Paszko indices. V. champinii and V. girdiana were apart from the other species by CVcl and CVci graphic representation and also formed a group apart in the dendrogram based on Euclidian distances. The chromosome pair number 3 harbors the secondary constriction and a satellite segment in all species analyzed with Giemsa staining and it may be the same observed after NOR banding technique. It seems that the process of speciation in the North American Euvitis species studied involved some discrete changes in chromosome morphometry which have been reflected in the asymmetry index.

The use of the Cytogenetic to Identify Mechanisms of Action of an Azo Dye in Allium Cepa Meristematic Cells

In the present study, cytotoxic, genotoxic and mutagenic actions of different concentrations (1, 10, 100 and 1000 μg/L) of an azo dye (BDCP – Black Dye Commercial Product) were evaluated using different cytogenetic techniques [(conventional dye, C banding, NOR banding, base-specific fluorochrome banding and fluorescent in situ hybridization (FISH)] applied to the Allium cepa test-organism. The use of conventional cytogenetic staining allowed us to determine that the azo dye induced cell death, chromosomal aberrations, nuclear alteration and micronuclei. By means of chromosome bandings and the FISH technique, it was possible to notice both cell and nucleolar alterations induced by BDCP, whose effects, in a general way, were not specific for any chromosome sites. The abnormalities observed made it possible to infer both aneugenic and clastogenic actions caused by the dye analyzed. The cytotoxic, genotoxic and mutagenic effects of BDCP were not completely eliminated, even after the recovery treatment of the A. cepa roots in water, showing a cumulative potential effect of the referred dye. Due to these results, we could infer that the BDCP may be a dangerous contaminant to the environment and, consequently, to the lives of the organisms exposed to it.

Molecular and Cytogenetic Evaluation of Four Indian Breeds of Sheep

Kumar, P., Kumar, K.G., Bhattacharya, T.K., Bhushan, B., Sharma, A. and Ahlawat, S.P.S. 2009. Molecular and cytogenetic evaluation of four Indian breeds of sheep. J. Appl. Anim. Res., 36: 261–266. This study was conducted to evaluate four breeds of Indian sheep namely Marwari, Mandya, Madras Red and Muzaffarnagari at cytogenetic and molecular genetic levels. The Karyotype revealed 54 numbers of chromosomes in both the sexes. The NORs were found on the telomeric ends of chromosomal pairs 1, 2, 3, 4 and 5. C-bands were found on all the autosomes while sex chromosomes were C-band negative. The G-bands were found on all metaphase chromosomes. The RAPD experiments produced 6 to 15 fingerprints with size ranging from 0.3 to 1.9 kb. The intrabreed homogeneity estimated in terms of band sharing frequency (BSF) was highest (0.95) in Marwari breed, whereas, lowest magnitude (0.90) was observed in Muzaffarnagari breed. The genetic distance based on MAPD was highest (9.5) between Marwari and Muzaffarnagari, whereas, the lowest magnitude (4.9) was found between Mandya and Muzaffarnagari breeds. It may be concluded that all four breeds of sheep studied have similarity in chromosomal configuration based on C, G and NOR banding patterns. The results will help in characterization and ultimately conservation of these sheep breeds.

Chromosome Characterization and Cytotaxonomic considerations on Characidae, Acestrorhynchidae and Cynodontidae (Pisces, Characiformes) from the Parana River (Argentina).

Abstract Characiformes have several families, being Characidae important due to the number and diversity of species. The present work presents the first cytogenetic characterization of Acestrorhynchus pantaneiro, Roeboides bonariensis, Cynopotamus kinkaidi, Galeocharax humeralis and Raphiodon vulpinus from Argentina (Parana river- Posadas, Misiones) stating some considerations on the cytotaxonomic relationships among them. Chromosome preparations were obtained from kidney cells by means of direct and cell culture methods. Chromosomes were stained conventionally and with C and NOR banding. Chromosome numbers ranged from 2n=50 (Acestrorhynchus pantaneiro, Acestrorhynchidae) and 2n=52 (Roeboides bonariensis, Cynopotamus kinkaidi and Galeocharax humeralis, Characidae) to 2n=54 (Raphiodon vulpinus, Cynodontidae) with different proportions among the chromosome types M-SM/ ST-A. All species show pale heterochromatin, however, some centromeric, telomeric and interstitial regions appear strongly stained. NOR regio...

Cytogenetics of two subspecies of White-Tailed deer (Odocoileus) from Venezuela

We studied the cytogenetics of two subspecies of deer in Venezuela: Odocoileus virginianus gymnotis, distributed almost nationwide and O. v. margaritae, endemic to Margarita Island. All individuals examined showed karyotypes of (2n=70), with the same fundamental number (FN=70). The first pair of chromosomes was metacen- tric (small-sized) and the other 33 pairs were acrocentric (medium- and small-sized). Chromosomes of sexual pair were metacentric (medium-sized (X) and small-sized (Y). C-Banding revealed pericentromeric heterochromatin in all the acrocentric chromosomes, while the metacentric autosomal pair did not show well defined bands. In both subspecies, the X chromosome presented pericentromeric constitutive heterochromatin. The Y chromosome did not show the presence of any C band in O. v. gymnotis. This could not be determined in O. v. margaritae because the only animal available was a female. NOR bands were located on the telomeric region of both chromosomes of pair 2 and in the centromeric region of pair 3. G-banding revealed similar patterns of bands in all chromosomes of both subspecies, except in the autosomal second pair. Odocoileus v. gymnotis showed a terminal band that seems to be absent in O. v. margaritae. These karyotypic analyses do not allow us to assure that the studied entities are two different species. We propose a series of chromosomal rearrangements the could have produced the current karyotypic diversity in the family Cervidae

The karyotypes of the thorny catfishes Wertheimeria maculata Steindachner, 1877 and Hassar wilderi Kindle, 1895 (Siluriformes: Doradidae) and their relevance in doradids chromosomal evolution

We studied the karyotypes of two doradids, the rare and endangered Wertheimeria maculata and a derived Amazonian species, Hassar wilderi. Cytogenetic characterization was assessed using conventional staining (Giemsa), C-banding, and NOR banding. Both species had 2n = 58 chromosomes but differed in their chromosome formulae, 24 m + 14sm + 8st + 12a for W. maculata and 32 m + 16sm + 10st for H. wilderi. In W. maculata heterochromatin was mainly telomeric, and three chromosomes had a fully heterochromatic arm; in H. wilderi heterochromatin was also predominantly telomeric and evident in many more chromosomes. Hassar wilderi also presented one pair of homologues with a fully heterochromatic arm. In both species, nucleolar organizer regions were restricted to one pair of subtelocentric chromosomes. Assuming a basal position for W. maculata, we hypothesized that underlying conserved diploid and NOR-bearing chromosome numbers, chromosomal evolution in doradids has involved pericentric inversions and an increase of heterochromatic blocks.