Interstitial Telomeric Signals Research Articles

A step forward in the genome characterization of the sugarcane borer, Diatraea saccharalis: karyotype analysis, sex chromosome system and repetitive DNAs through a cytogenomic approach.

Moths of the family Crambidae include a number of pests that cause economic losses to agricultural crops. Despite their economic importance, little is known about their genome architecture and chromosome evolution. Here, we characterized the chromosomes and repetitive DNA of the sugarcane borer Diatraea saccharalis using a combination of low-pass genome sequencing, bioinformatics, and cytogenetic methods, focusing on the sex chromosomes. Diploid chromosome numbers differed between the sexes, i.e., 2n = 33 in females and 2n = 34 in males. This difference was caused by the occurrence of a WZ1Z2 trivalent in female meiosis, indicating a multiple sex-chromosome system WZ1Z2/Z1Z1Z2Z2. A strong interstitial telomeric signal was observed on the W chromosome, indicating a fusion of the ancestral W chromosome with an autosome. Among repetitive DNAs, transposable elements (TEs) accounted for 39.18% (males) to 41.35% (females), while satDNAs accounted for only 0.214% (males) and 0.215% (females) of the genome. FISH mapping revealed different chromosomal organization of satDNAs, such as single localized clusters, spread repeats, and non-clustered repeats. Two TEs mapped by FISH were scattered. Although we found a slight enrichment of some satDNAs in the female genome, they were not differentially enriched on the W chromosome. However, we found enriched FISH signals for TEs on the W chromosome, suggesting their involvement in W chromosome degeneration and differentiation. These data shed light on karyotype and repetitive DNA dynamics due to multiple chromosome fusions in D. saccharalis, contribute to the understanding of genome structure in Lepidoptera and are important for future genomic studies.

Extensive genomic reshuffling involved in the karyotype evolution of genus Cerradomys (Rodentia: Sigmodontinae: Oryzomyini).

Rodents of the genus Cerradomys belong to the tribe Oryzomyini and present high chromosome variability with diploid numbers ranging from 2n=46 to 60. Classical cytogenetics and fluorescence in situ hybridization (FISH) with telomeric and whole chromosome-specific probes of another Oryzomyini, Oligoryzomys moojeni (OMO), were used to assess the karyotype evolution of the genus. Results were integrated into a molecular phylogeny to infer the hypothetical direction of chromosome changes. The telomeric FISH showed signals in telomeres in species that diverged early in the phylogeny, plus interstitial telomeric signals (ITS) in some species from the most derived clades (C. langguthi, C. vivoi, C. goytaca, and C. subflavus). Chromosome painting revealed homology from 23 segments of C. maracajuensis and C. marinhus to 32 of C. vivoi. Extensive chromosome reorganization was responsible for karyotypic differences in closely related species. Major drivers for genomic reshuffling were in tandem and centric fusion, fission, paracentric and pericentric inversions or centromere repositioning. Chromosome evolution was associated with an increase and decrease in diploid number in different lineages and ITS indicate remnants of ancient telomeres. Cytogenetics results corroborates that C. goytaca is not a junior synonym of C. subflavus since the karyotypic differences found may lead to reproductive isolation.

Cytogenetic characterization and description of an X1X1X2X2/X1X2Y sex chromosome system in Collichthys lucidus (Richardson, 1844)

The chromosomes of spinyhead croaker Collichthys lucidus (Richardson, 1844) were characterized for the first time by fluorescence staining, self genomic in situ hybridization (self-GISH), and multicolor fluorescence in situ hybridization (FISH) with 18S rDNA, 5S rDNA and telomeric sequence probes. The female karyotype has exclusively 24 pairs of acrocentric chromosomes (2n=48a, NF=48), while the male one consists of 22 pairs of acrocentric chromosomes, 2 monosomic acrocentric chromosomes and a metacentric chromosome (2n=1m+46a, NF=48). The difference between female and male karyotypes indicates the presence of a sex chromosome of X1X1X2X2/X1X2Y type, where Y is the unique metacentric chromosome in the male karyotype. As revealed by FISH, 5S rDNA and 18S rDNA sites were mapped at syntenic position of the largest acrocentric chromosome (X1), and the short arms of the Y chromosome as well. An X1-chromosome specific interstitial telomeric signal (ITS) was detected overlapping the 5S rDNA sites. In addition, self-GISH revealed that the repetitive DNAs accumulated on all the putative sex chromosome. Chromosome fusion accompanied by a partial deletion in the ancestral karyotype (2n=48a) is hypothesized for the origin of such multiple sex chromosome system. The present study, as the first description of differentiated sex chromosome in family Sciaenidae, will give clues to the studies on the sex chromosome of other Sciaenids.

Evolution of Karyotypes in Chameleons.

The reconstruction of the evolutionary dynamics of karyotypes and sex determining systems in squamate reptiles is precluded by the lack of data in many groups including most chameleons (Squamata: Acrodonta: Chamaeleonidae). We performed cytogenetic analysis in 16 species of chameleons from 8 genera covering the phylogenetic diversity of the family and also phylogenetic reconstruction of karyotype evolution in this group. In comparison to other squamates, chameleons demonstrate rather variable karyotypes, differing in chromosome number, morphology and presence of interstitial telomeric signal (ITS). On the other hand, the location of rDNA is quite conserved among chameleon species. Phylogenetic analysis combining our new results and previously published data tentatively suggests that the ancestral chromosome number for chameleons is 2n = 36, which is the same as assumed for other lineages of the clade Iguania, i.e., agamids and iguanas. In general, we observed a tendency for the reduction of chromosome number during the evolution of chameleons, however, in Rieppeleon brevicaudatus, we uncovered a chromosome number of 2n = 62, very unusual among squamates, originating from a number of chromosome splits. Despite the presence of the highly differentiated ZZ/ZW sex chromosomes in the genus Furcifer, we did not detect any unequivocal sexual differences in the karyotypes of any other studied species of chameleons tested using differential staining and comparative genomic hybridization, suggesting that sex chromosomes in most chameleons are only poorly differentiated.

Combining FISH and model-based predictions to understand chromosome evolution in Typhonium (Araceae)

Since the advent of molecular phylogenetics, numerous attempts have been made to infer the evolutionary trajectories of chromosome numbers on DNA phylogenies. Ideally, such inferences should be evaluated against cytogenetic data. Towards this goal, we carried out phylogenetic modelling of chromosome number change and fluorescence in situ hybridization (FISH) in a medium sized genus of Araceae to elucidate if data from chromosomal markers would support maximum likelihood-inferred changes in chromosome numbers among close relatives. Typhonium, the focal genus, includes species with 2n = 65 and 2n = 8, the lowest known count in the family. A phylogeny from nuclear and plastid sequences (96 taxa, 4252 nucleotides) and counts for all included species (15 of them first reported here) were used to model chromosome number evolution, assuming discrete events, such as polyploidization and descending or ascending dysploidy, occurring at different rates. FISH with three probes (5S rDNA, 45S rDNA and Arabidopsis-like telomeres) was performed on ten species with 2n = 8 to 2n = 24. The best-fitting models assume numerous past chromosome number reductions. Of the species analysed with FISH, the two with the lowest chromosome numbers contained interstitial telomeric signals (Its), which together with the phylogeny and modelling indicates decreasing dysploidy as an explanation for the low numbers. A model-inferred polyploidization in another species is matched by an increase in rDNA sites. The combination of a densely sampled phylogeny, ancestral state modelling and FISH revealed that the species with n = 4 is highly derived, with the FISH data pointing to a Robertsonian fusion-like chromosome rearrangement in the ancestor of this species.

Karyotype and Chromosome Variability in the Armadillo Chaetophractus villosus in Argentina

Karyotype and cytotype variations for the large hairy armadillo (Chaetophractus villosus) were studied throughout the species' Argentine distribution. Peripheral blood lymphocyte cultures of 421 animals were used to obtain mitotic metaphases. Preparations were subjected to conventional staining, G- and C-banding, and FISH involving a telomeric probe. Meiotic analysis was performed on testis material from 10 adults. Spermatocytes were examined for synaptonemal complexes in microspreads. The karyotype (2n = 60 XX/XY; FN = 84 without XY) showed an autosomal complement of 6 metacentric and 7 submetacentric chromosomes; the remainder was acrocentric. The X chromosome was submetacentric and the Y acrocentric. Centromeric C+ marks were observed in all chromosomes except pair 16. Three NOR signals were detected in 6q, 12p, and 26p. Two chromosomal rearrangements were characterized in chromosome pair 1 a pericentric inversion seen in the material from Jacinto Aráuz, General Madariaga and Pellegrini and a deletion in the material from Loma Verde. Interstitial telomeric signals were observed in chromosome pairs 4, 12, 16, and 26. Pachytene spermatocyte analysis confirmed the basic chromosome number and morphologies observed in mitotic karyotypes. The evolution of C. villosus involved chromosomal rearrangements as recorded for other species of its superorder. The present results establish the basis for the cytogenetic characterization of this species.

Telomeres in Fishes

In fishes, as in other vertebrate species, the DNA component of the telomeres consists of the tandemly repeated TTAGGG motif. The length of the telomeric arrays in fishes ranges from 2 to 25 kb and shortens with age in some of the species. To date, chromosomal distribution of the telomeric DNA sequences has been examined in approximately 80 fish species of which about 42% show additional telomeric hybridization signals far from the chromosomal termini. Based on the chromosomal location, such internally located telomeric repeats may be classified into 4 categories: (1) telomeric DNA sequences located at the pericentromeric regions, (2) interstitial telomeric sites observed between centromeres and the bona fide telomeres, (3) telomeric DNA sequences that scatter along the nucleolus organizer regions, and (4) telomeric DNA repeats interspersed with the entire chromosomes. Most of the pericentromeric and interstitial telomeric sequences in fish are possible relicts of chromosome fusion events. The origin of the telomeric sequences co- localizing with the major rDNA sequences or scattered along the whole chromosomes is not clear. Internally located telomeric repeats are considered as ‘hot spots' for recombination and thus may potentially increase the rates of chromosome breaks and rearrangements leading to the various chromosomal polymorphisms in fishes. FISH with telomeric probe applied to metaphase spreads of androgenetic specimens that hatched from eggs exposed to ionizing radiation before insemination enabled the detection of small radiation-induced fragments of maternal chromosomes. Remnants of the irradiated chromosomes were found to be ring chromosomes with the interstitial telomeric signals, telomerless rings, fragments with fused sister chromatids, and linear fragments with telomeres detected at both of their ends. The increasing availability of techniques enabling the study of fish telomeres and telomerase and the easy access to numerous fish species strongly confirm that these animals are promising models in research concerning the role of telomeres and telomerase in vertebrate aging, repair of ionizing radiation-induced DNA double strand breaks, and chromosomal rearrangements.

Identification of Telomeric Sequences in Pigs with Rearranged Karyotype Using Prins Technique / Identyfikacja sekwencji telomerowych u świń z rearanżacją kariotypu przy wykorzystaniu techniki PRINS

Abstract Using PRINS technique with oligonucleotide primer complementary to the telomere sequence repeats (TTAGGG )n, the interstitial telomeric signals in 6q22 and 7q13 unstable chromosome regions in pigs with t(7;13)(q13;q46) reciprocal translocation were identified. The intrachromosomal localization of telomeric repeats specific for 7q13 region were revealed to strictly correspond to breakpoint of this translocation as well as several others described earlier. The present study showed the usefulness of PRINS technique for identification of intrachromosomal telomeric sequences in pig genome regions, extremely prone to breakage and predisposed to chromosome reorganizations, especially with regard to the clinical cases of karyotype defects.

First cytogenetic information for Drymoreomys albimaculatus (Rodentia, Cricetidae), a recently described genus from Brazilian Atlantic Forest

The recently described taxon Drymoreomys albimaculatus is endemic to the Brazilian Atlantic Forest and its biology and genetics are still poorly known. Herein, we present, for the first time, the karyotype of the species using classical and molecular cytogenetics, which showed 2n=62, FN=62, and interstitial telomeric signals at the sex chromosomes. Nuclear and mitochondrial DNA sequences from the two karyotyped individuals verify the taxonomic identity as the recently described Drymoreomys albimaculatus and confirm the relationship of the species with other Oryzomyini. Additionally, external morphological information is provided.

Molecular Cytogenetics (FISH, GISH) of Coccinia grandis: A ca. 3 myr-Old Species of Cucurbitaceae with the Largest Y/Autosome Divergence in Flowering Plants

The independent evolution of heteromorphic sex chromosomes in 19 species from 4 families of flowering plants permits studying X/Y divergence after the initial recombination suppression. Here, we document autosome/Y divergence in the tropical Cucurbitaceae Coccinia grandis, which is ca. 3 myr old. Karyotyping and C-value measurements show that the C. grandis Y chromosome has twice the size of any of the other chromosomes, with a male/female C-value difference of 0.094 pg or 10% of the total genome. FISH staining revealed 5S and 45S rDNA sites on autosomes but not on the Y chromosome, making it unlikely that rDNA contributed to the elongation of the Y chromosome; recent end-to-end fusion also seems unlikely given the lack of interstitial telomeric signals. GISH with different concentrations of female blocking DNA detected a possible pseudo-autosomal region on the Y chromosome, and C-banding suggests that the entire Y chromosome in C. grandis is heterochromatic. During meiosis, there is an end-to-end connection between the X and the Y chromosome, but the X does not otherwise differ from the remaining chromosomes. These findings and a review of plants with heteromorphic sex chromosomes reveal no relationship between species age and degree of sex chromosome dimorphism. Its relatively small genome size (0.943 pg/2C in males), large Y chromosome, and phylogenetic proximity to the fully sequenced Cucumis sativus make C. grandis a promising model to study sex chromosome evolution.

Genomic Instability and Telomere Fusion of Canine Osteosarcoma Cells

Canine osteosarcoma (OSA) is known to present with highly variable and chaotic karyotypes, including hypodiploidy, hyperdiploidy, and increased numbers of metacentric chromosomes. The spectrum of genomic instabilities in canine OSA has significantly augmented the difficulty in clearly defining the biological and clinical significance of the observed cytogenetic abnormalities. In this study, eight canine OSA cell lines were used to investigate telomere fusions by fluorescence in situ hybridization (FISH) using a peptide nucleotide acid probe. We characterized each cell line by classical cytogenetic studies and cellular phenotypes including telomere associated factors and then evaluated correlations from this data. All eight canine OSA cell lines displayed increased abnormal metacentric chromosomes and exhibited numerous telomere fusions and interstitial telomeric signals. Also, as evidence of unstable telomeres, colocalization of γ-H2AX and telomere signals in interphase cells was observed. Each cell line was characterized by a combination of data representing cellular doubling time, DNA content, chromosome number, metacentric chromosome frequency, telomere signal level, cellular radiosensitivity, and DNA-PKcs protein expression level. We have also studied primary cultures from 10 spontaneous canine OSAs. Based on the observation of telomere aberrations in those primary cell cultures, we are reasonably certain that our observations in cell lines are not an artifact of prolonged culture. A correlation between telomere fusions and the other characteristics analyzed in our study could not be identified. However, it is important to note that all of the canine OSA samples exhibiting telomere fusion utilized in our study were telomerase positive. Pending further research regarding telomerase negative canine OSA cell lines, our findings may suggest telomere fusions can potentially serve as a novel marker for canine OSA.

Distribution of Telomeric DNA Sequences on the X-Radiation-Induced Chromosome Fragments Observed in the Genome of Androgenetic Brook Trout (Salvelinus fontinalis, Mitchill 1814)

Cytogenetic screening of the androgenetic brook trout (Salvelinus fontinalis, Mitchill 1814) offspring hatched from eggs exposed to 420 Gy of X-radiation before insemination exhibited residues of the irradiated maternal nuclear genome in the form of small chromosome fragments. Remnants of the irradiated chromosomes had different sizes, and their number varied intraindividually from 1 to 15. To efficiently pass through the series of the cell divisions, such chromosome fragments must have had functional kinetochores. Distribution patterns of the telomeric hybridization signals on the chromosome fragments enabled us to distinguish their 3 groups: (i) telomere-less ring chromosomes with fused broken chromosome arms, (ii) rings formed in the course of fusion of the radiation-broken chromosome arm with the opposite telomeric region and exhibiting interstitial telomeric signals at the fusion point, and (iii) chromosome fragments with fused unprotected sister chromatids of 1 broken arm and intact telomeres from the other arm. Disturbances during segregation of such fragments, mainly breakages during anaphase, may partially explain intraindividual variation in the number and size of the chromosome fragments observed in the androgenetic brook trout.

Duplicated chromosomal fragments stabilize shortened telomeres in normal human IMR‐90 cells before transition to senescence

To assess why during in vitro aging of fibroblasts the maintenance of chromosomal stability is effective or occasionally fails, a detailed cytogenetic analysis was performed in normal human IMR-90 fetal lung fibroblasts. The onset of senescence was inferred from proliferation activity, expression pattern of cell cycle regulating proteins, activity of β-galactosidase, and morphological features. Over the period of proliferation, a moderate increase of non-transmissible structural chromosomal aberrations was observed. In addition, using fluorescence in situ hybridization (mFISH and mBAND) techniques, we detected clonally expanding translocations in up to 70% of the analyzed metaphases, all involving one homolog of chromosome 9 as an acceptor. Notably, chromosomes are randomly involved as donor-chromosomes of the translocated terminal acentric fragments. These fragments result from duplication because the donor chromosomes are apparently unchanged. Interstitial telomeric signals were detectable at fusion sites, most likely belonging to chromosome 9. Quantitative fluorescence in situ hybridization (QFISH) detecting telomere sequences, followed by mFISH technique revealed that already in young cells the respective telomeres of one chromosome 9 were particularly short. For the first time, we have observed dysfunctional telomeres of one specific chromosome in normal human cells that have been stabilized by duplicated terminal sequences.

Karyotypic Evolution of Hapalomys Inferred from Chromosome Painting: A Detailed Characterization Contributing New Insights into the Ancestral Murinae Karyotype

We report on the construction of a comparative chromosome map between the emblematic laboratory rat, Rattus norvegicus (RNO), and Delacour’s Marmoset rat, Hapalomys delacouri (HDE), based on cross-species fluorescence in situ hybridization with R. norvegicus painting probes. Sixteen R. norvegicus chromosomes (RNO 3–6, 8, 10–15, 17–20, and X) were retained in their entirety (as a conserved block or as a single chromosome) in the H. delacouri genome. The remaining 5 R. norvegicus chromosomes (RNO 1, 2, 7, 9, and 16) produced 2 signals in the H. delacouri karyotype. Our analysis allowed the detection of an X-autosome translocation between RNO X and 11 that occurred convergently in an unrelated species, Bandicota savilei, and a single B chromosome that accounts for the 2n = 48 karyotype observed in this specimen. In total, the rat chromosome paints revealed 27 segments of conserved synteny in H. delacouri. The analysis showed 7 NOR bearing pairs in H. delacouri (HDE 1, 3, 6, 7, 8, 10, and 13) and the occurrence of an interstitial telomeric signal at the centromeric regions of 8 H. delacouri chromosomes (HDE 3, 10, 11, 12, 13, 16, 19, and 22). These data, together with published comparative maps, enabled a revision of the previously postulated murine ancestral condition suggesting that it probably comprised a wholly acrocentric karyotype with 2n = 46–50.

한국 재래닭 염색체의 텔로미어 분포 양상

텔로미어(telomere)는 염색체 양 말단에 위치하는 DNA와 단백질의 복합체로서 (TTAGGG)n의 단순 반복 염기 서열로 이루어져 있다. 그러나 일부 조류 및 척추동물의 경우 염색체의 양 말단 부위외 간질적 위치에도 telomeric DNA sequence가 분포한다. 본 연구는 닭 염색체에 있어 telomeric DNA의 분포 양상을 제시하고자 한국 재래닭의 초기 배아로부터 염색체 표본을 제작하고, telomeric DNA probe를 이용한 FISH를 수행하여 염색체 상 텔로미어의 분포 양상을 분석하였다. 분석 결과, 닭의 모든 염색체 양 말단부에 텔로미어가 분포하는 것으로 나타났으며, 더불어 대형 염색체 중 1번의 1q32, 1p11, 1p23 위치와 2번 염색체의 2q24 및 3번 염색체 3q32에 interstitial telomeric signal(ITS)이 존재하는 것이 확인되었다. 이러한 한국 재래닭 염색체의 텔로미어 분포 양상은 이전 Gallus domesticus에서 발표한 분포 양상과 거의 일치한 것으로 나타났다. 한국 재래닭의 각 염색체별 텔로미어 함유율은 4.6~16.3% 정도로 분석되었으며, 거의 대부분의 염색체에서 단완 말단부의 telomeric DNA의 함량이 장완 말단부보다 높은 것으로 나타났다. 닭 염색체에서 ITS의 존재와 분포 양상은 핵형학적으로 진화 과정 중 염색체 간의 융합에 의해 신생 염색체가 형성되었을 가능성을 시사한다. Telomeres are nucleoprotein structures at the ends of chromosomes consisting of DNA sequences arranged in tandemly repeated units <TEX>$(TTAGGG)_n$</TEX>. However, this hexamers can also occur at non-telomeric sites in some avians and vertbrate. This study was carried out to present the distribution of telomeric DNA sequences in Korean Native Chicken chromosomes. The fluorescence in situ hybridization technique using a telomeric DNA probe was performed to the metaphase spreads of chicken early embryonic cells. Telomeric DNA signals were detected at the ends of chromosomes including macrochromosomes and microchromosomes. In chicken, surprisingly, chromosome 1 showed very distinct interstitial telomeric DNA hybridization patterns which located two interstitial sites in the p-arm at 1p11 and 1p23, and one in the q-arm at 1q32. In chromosome number 2 and 3 also displayed interstitial telomeric signals (ITS) in the long arms at 2q24 and 3q32, respectively. The pattern of telomeric DNA distribution in Korean Native Chicken chromosomes was in agreement with a previously reported in Gallus domesticus. The relative amount of telomeric DNA sequences in each macrochromosomes ranged from 4.6% to 16.3%. Distribution of telomeric DNAs at the end of p-arm was much more than that of q-arm in almost chicken chromosomes. The distribution of ITS in chicken chromsomes implicate to tandem chromosome fusions that might have occurred during the process of karyotype evolution.

Chromosomal mapping of rDNAs, core histone genes and telomeric sequences in Perumytilus purpuratus (Bivalvia: Mytilidae)

The chromosomes of the mussel Perumytilus purpuratus were analyzed by means of 4’,6-diamidino-2-phenylindole (DAPI)/propidium iodide (PI) and chromomycin A3 (CMA)/DAPI fluorescence staining and fluorescent in situ hybridization (FISH) with major rDNA, 5S rDNA, core histone gene and telomeric probes. The diploid chromosome number in this species is 32. The karyotype is composed by two metacentric, five submetacentric, one subtelo/submetacentric and eight subtelocentric chromosome pairs. The nucleolar organizing regions (NORs) detected by FISH using major rDNA probes were terminally located on the long arms of two pairs of chromosomes. NORs were associated with bright CMA fluorescence and dull DAPI fluorescence but not all the NORs showed bright CMA fluorescence in a given cell; intra- and inter-individual variability was found in this character. FISH detected a similar variability on the number of major rDNA signals. This species shows three 5S rDNA clusters on two chromosome pairs. Two of the clusters are terminal and intercalary located on one of the two NOR-bearing chromosome pairs; the third cluster is on the long arm of another chromosome pair. Core histone genes are clustered on a single locus near the centromere on the long arm of a different chromosome pair. This locus is not condensed in prophase I cells therefore suggesting a certain degree of histone gene expression during meiosis. As expected, P. purpuratus shows telomeric signals at both ends of every single chromosome but interstitial telomeric signals were also detected at centromeric positions on two chromosome pairs.

Karyotypic characterization of Trachemys dorbigni (Testudines: Emydidae) and Chelonoidis (Geochelone) donosobarrosi (Testudines: Testudinidae), two species of Cryptodiran turtles from Argentina

We describe for the first time the karyotypes of two species of Cryptodiran turtles from Argentina, namely, Trachemys dorbigni (Emydidae) and Chelonoidis (Geochelone) donosobarrosi (Testudinidae). The karyotype of T. dorbigni (2n = 50) consists of 13 pairs of macrochromosomes and 12 pairs of microchromosomes, whereas the karyotype of C. donosobarrosi (2n = 52) consists of 11 pairs of macrochromosomes and 15 pairs of microchromosomes. Fluorescence in situ hybridization (FISH) with a (TTAGGG)n telomeric probe showed that the chromosomes of these species have four telomeric signals, two at each end, indicating that none of the chromosomes of T. dorbigni and C. donosobarrosi are telocentric. The fact that no interstitial telomeric signals were observed after FISH, suggests that interstitial telomeric sequences did not have a major role in the chromosomal evolution of these species. Additional data will be needed to elucidate if interstitial telomeric sequences have a major role in the karyotypic evolution of Testudines.

Application of primed in situ DNA synthesis (PRINS) with telomere human commercial kit in molecular cytogenetics of Equus caballus and Sus scrofa scrofa.

Recently, molecular techniques have become an indispensable tools for cytogenetic research. Especially, development of in situ techniques made possible detection at the chromosomal level, genes as well as repetitive sequences like telomeres or the DNA component of telomeres. One of these methods is primed in situ DNA synthesis (PRINS) using an oligonucleotide primer complementary to the specific DNA sequence. In this report we described application of PRINS technique with telomere human commercial kit to telomere sequences identification. This commercial kit may be use to visualization of interstitial telomeric signal in pig genome. PRINS is attractive complement to FISH for detection of DNA repetitive sequences and displays lower level of non-specific hybridization than conventional FISH.

The origin of an unusual sex chromosome constitution in Acomys sp. (Rodentia, Muridae) from Tanzania

This paper describes a case which presents an evident variation from the "standard" XX/XY sex chromosomal constitution in a rodent, Acomys sp. This species known to be found in three localities of East Africa has only recently been separated from A. spinosissimus, its closest relative. In our study, five specimens of Acomys sp. and eight specimens of A. spinosissimus were live-trapped in five localities. Comparisons between the two taxa assed by G-banding show a complete homology in the chromosomal shape and banding pattern for 29 pairs of chromosomes corresponding to the complete autosomal set of A. spinosissimus. However, while all the A. spinosissimus analysed have 2n = 60 and a XY-XX system, in Acomys sp. males and females constitute mosaics for sex chromosomes in the bone marrow cells. Females (2n = 59, 60) have an excess (97%) of aneuploid cells with one single giant X chromosome, and males (2n = 60, 61) show X0/XY cells occurring in somatic tissues and XY cells in the germinal lineage. In addition, an odd heterochromatic submetacentric chromosome was identified in all the cells examined in two males and a female of Acomys sp. Since this chromosome was not related to sex determination and it is not present in all the analysed specimens, it can be considered as a B chromosome. Finally, the in situ fluorescence hybridisation (FISH) with telomeric probes showed a very intense interstitial telomeric signal (ITS) at the medial part on the long heterochromatic arm of the X chromosome. This could be due to recent chromosomal rearrangement.

소, 돼지 염색체의 telomeric DNA 분포 양상

Telomeres are nucleoprotein structures at the ends of chromosomes consisting of tandem repeat sequences of . (TTAGGG)n. Telomeres serve as guardians of the genome, protect individual chromosomes within the nucleus, and help in meiotic pairing of homologous chromosomes. To investigate the telomere distributions of cattle and pig chromosomes, fluorescence in situ hybridization(FISH) was carried out on metaphase spreads of in vitro fibroblast cultures from Holstein and Landrace using a human telomeric DNA repeat probe. Results indicate that the distinct double spots on both ends of chromosomes of cattle and pigs were observed. In cattle, there was a random variation in the intensity of telomere signals among chromosomes. In pigs, an interstitial telomeric signal was observed on the chromosome 6q1 of all the cells examined. According to quantitative fluorescence in situ hybridization(Q-FISH) analysis, some chromosomes had consistently much more telorneres at one end of chromosomes. In general, both species had consistently much more telomeres at q-end than p-end on most of chromosomes. The relative amount of telomeres on bovine chromosomes was higher than that on pig chromosomes. In additions, Y chromosome had the highest relative amount of telorneres in cattle and pigs.