5S rDNA Signals Research Articles

Multicolor FISH와 Feulgen 염색법을 이용한 Angelica속 식물의 세포유전학적 분석

Karyotype analysis and chromosomal localization of 5S and 45S rDNAs using multi-color fluorescence in situ hybridization (McFISH) technique were carried out in two Angelica species. The numbers of diploid chromosomes were the same in two same in two species as 2n=22, however the lengths of chromosomes were varied from 4.25 to 6.50 <TEX>${\mu}{\textrm}{m}$</TEX> in A gigas and 4.95 to 8.50 <TEX>${\mu}{\textrm}{m}$</TEX> in A acutiloba. The chromosomes of A. gigas were composed of five metacentric and six submetacentric pairs, while those of A. acutiloba were six metacentic, one submetacentric and four subtelocentric paris. In FISH experiments, the numbers and size of 45S rDNA signals were varied between two species, however dach signal of the 5S rDNA was observed in two species.

2배체 담배 Nicotiana plumbaginifolia의 핵형 분석과 rDNAs의 Physical Mapping

2배체 담배인 Nicotiana plumbaginifolia를 대상으로 상염색법과 FISH 기법을 통한 염색체 분석을 수행하여 다음과 같은 결과를 얻었다. N. plumbaginifolia의 체세포 염색체 수는 2n=20이며, arm ratio 비교를 통한 핵형 분석에서 중기 염색체 조성은 3쌍의 중부 염색체 (염색체 1,2 및 7)와 7쌍의 차중부 염색체 (염색체 3, 4, 5, 6, 7, 9 및 10)로 관찰되었다. 염색체의 길이는 2.29~4.50 <TEX>$\mu\textrm{m}$</TEX>로 나타났으며, 염색체 1번과 2번은 부수체 염색체로 관찰되었다. 5S와 45S rDNA를 탐침으로 FISH를 수행한 결과 2번 염색체의 동원체 부위에서 한 쌍의 5S signal이 확인되었고, 1번 염색체의 부수체에서 한 쌍의 45S signal이 관찰되었다. Karyotype analysis and chromosomal localization of 5S and 45S rDNAs using FISH (fluorescence in situ hybridization) technique were carried out in Nicotiana plumbaginifolia. Somatic chromosome number of N. plumbaginifolia was 2n=20 and kyarotype was composed of three pairs of metacentric chromosomes (1,2 and 7) and seven pairs of submetacentric chromosomes (3, 4, 5, 6, 8, 9 and 10). Length of chromosomes was ranged from 2.29 to 4.50 <TEX>${\mu}{\textrm}{m}$</TEX>. Chromosome 1 and 2 were identified as satellite chromosomes. In FISH experiment, one pair of 5S rDNA signals was detected on the centromeric region of chromosome 2 and one pair of 45S rDNA signals was detected on the terminal region of chromosome 1.

Karyotype analysis ofLilium longiflorumandLilium rubellumby chromosome banding and fluorescence in situ hybridisation

Detailed karyotypes of Lilium longiflorum and L. rubellum were constructed on the basis of chromosome arm lengths, C-banding, AgNO3 staining, and PI-DAPI banding, together with fluorescence in situ hybridisation (FISH) with the 5S and 45S rDNA sequences as probes. The C-banding patterns that were obtained with the standard BSG technique revealed only few minor bands on heterologous positions of the L. longiflorum and L. rubellum chromosomes. FISH of the 5S and 45S rDNA probes on L. longiflorum metaphase complements showed overlapping signals at proximal positions of the short arms of chromosomes 4 and 7, a single 5S rDNA signal on the secondary constriction of chromosome 3, and one 45S rDNA signal adjacent to the 5S rDNA signal on the subdistal part of the long arm of chromosome 3. In L. rubellum, we observed co-localisation of the 5S and 45S rDNA sequences on the short arm of chromosomes 2 and 4 and on the long arms of chromosomes 2 and 3, and two adjacent bands on chromosome 12. Silver staining (Ag-NOR) of the nucleoli and NORs in L. longiflorum and L. rubellum yielded a highly variable number of signals in interphase nuclei and only a few faint silver deposits on the NORs of mitotic metaphase chromosomes. In preparations stained with PI and DAPI, we observed both red- and blue-fluorescing bands at different positions on the L. longiflorum and L. rubellum chromosomes. The red-fluorescing or so-called reverse PI-DAPI bands always coincided with rDNA sites, whereas the blue-fluorescing DAPI bands corresponded to C-bands. Based on these techniques, we could identify most of chromosomes of the L. longiflorum and L. rubellum karyotypes.

Chromosome phylogeny of Zamia and Ceratozamia by means of Robertsonian changes detected by fluorescence in situ hybridization (FISH) technique of rDNA

Appearance and location of 45S rDNA and 5S rDNA signals were compared in chromosomes of nine species of the aneuploid Zamia and their taxonomically and phylogenetically closely related Ceratozamia mexicana. The 45S rDNA signal was detected in the proximal region of six chromosomes in Zamia angustifolia, Z. integrifolia, Z. pumila and Z. pygmaea (all 2n=16); in the proximal region of 6–14 chromosomes in Z. furfuracea, Z. loddigesii, Z. skinneri and Z. vazquezii (all 2n=18); and on the proximal region of 20 chromosomes in Z. muricata (2n=23). The 5S rDNA signals were commonly seen near the terminal region of the short arm of two metacentric chromosomes in the four species with 2n=16 and Z. furfuracea, Z. loddigesii and Z. vazquezii with 2n=18. Other 5S rDNA signals were seen near the terminal region of two terminal-centromeric chromosomes in Z. skinneri and near the terminal region of a metacentric and a telocentric chromosomes in Z. muricata. In contrast, those with 45S and 5S rDNA signals were exhibited in chromosomes of Ceratozamia mexicana in a different manner from those in the nine species of Zamia; the 45S rDNA signal in the terminal region of four metacentric and two submetacentric chromosomes and the 5S rDNA signal near the proximal region of two metacentric chromosomes.

Phylogenetic Analysis and Karyotype Evolution in the Genus Clivia(Amaryllidaceae)

Phylogenetic relationships of five taxa of Clivia, one probable new species plus four recognized species, and three outgroup species were studied using sequences of the nuclear ribosomal 5S non-transcribed spacer and the internal transcribed spacer (ITS) of 45S rDNA. Analysis of the data sets separately generated some well-supported groupings and congruent phylogenies. Clivia miniata and C. gardenii are closely related. ‘Robust Gardenii’, the putative new species, is a sister clade of this group. Clivia nobilis is distantly related to these three taxa and C. caulescens occupies an intermediate position between the two groups. Chromosome locations and distribution patterns of the 5S nuclear ribosomal gene in the species of Clivia were investigated using fluorescencein situ hybridization (FISH). In all species, only one pair of 5S rDNA signals was observed. These were located on the short arm of chromosome 8, at the position of the interstitial C-bands. The phylogenies obtained from the DNA sequences together with the chromosome data accumulated here and previously published information on the location of the 45S rDNA sites have been used to postulate evolutionary trends in Clivia chromosomes.

Karyotype analysis of Nicotiana kawakamii Y. Ohashi using DAPI banding and rDNA FISH

Prometaphase cells were used to analyze the karyotype of Nicotiana kawakamii Y. Ohashi by means of sequential Giemsa/CMA/DAPI staining and multicolor fluorescence in situ hybridization with 5S and 18S rDNA. Observation of the DAPI-stained prometaphase spreads indicated that N. kawakamii had six pairs of large chromosomes, one pair of medium-sized chromosomes and five pairs of small chromosomes. The six pairs of large chromosomes possessed remarkable DAPI bands, and each could be identified from both the DAPI banding pattern and the length of the short arm. The DAPI banding pattern was approximately identical to the CMA and Giemsa banding patterns. Hybridization signals of the 18S rDNA probe were detected on two pairs of large chromosomes. In addition, two pairs of small chromosomes were identified based on the position of the 5S rDNA signals. An idiogram of N. kawakamii chromosomes was produced based on DAPI bands and rDNA loci.

Molecular cytogenetic identification of the parental genomes in the intergeneric hybrid between Leucanthemella linearis and Nipponanthemum nipponicum during meiosis and mitosis

Abstract The genomes of Leucanthemella linearis (2n=18) and Nipponanthemum nipponicum (2n=18) were studied in the chromosome complement of their F1 hybrid (2n=18) during meiosis and mitosis by DNA in situ hybridization and fluorochrome banding. Genomic in situ hybridization (GISH) using the total genomic DNA of N. nipponicum as a probe showed that the parental genomes in the chromosome complement of the hybrid were generally in separate domains during meiosis. Chromosome translocations were encountered and 18 univalents were usually observed at diakinesis and metaphase I. Meiotic abnormalities of chromosomes such as bridges, laggards and fragments were manifested mainly in the genome of L. linearis and after fluorescence in situ hybridization (FISH)using the 45S rDNA pTa71 probe showed that the parental genomes were differentiable in the hybrid. One 45S rDNA signal appeared on one of the nine chromosomes originating from N. nipponicum and thr e signals appeared in three of the nine chromosomes originating...

Identification and chromosomal location of tandemly repeated DNA sequences in Allium cepa.

A 314-bp tandemly repeated DNA sequence, named pAc074, was characterized in Allium cepa by fluorescence in situ hybridization (FISH) analyses using random amplified fragment as probe. The nucleotide sequences of the clone pAc074 is partially homologous to the satellite DNA sequences, ACSAT1, ACSAT2, and ACSAT3, of A. cepa with 81%, 81% and 78% similarity, respectively. Our sequential C-banding and FISH with pAc074 probe also clearly showed a close relation between Cheterochromatin at telomeric region and pAc074 sequences on all the chromosomes except on chromosome 6. On the long arm of chromosome 7, pAc074 sequences appeared as interstitial band which did not correspond to C-heterochromatin bands. Instead, the C-heterochromatin bands corresponded with the 5S rDNA signals. This is the first evidence of simultaneous banding of the 5S rDNA and C-band in A. cepa.

Molecular Cytogenetic Organization of 5S and 18S-26S rDNA Loci in White Clover (Trifolium repensL.) and Related Species

The genusTrifolium(Leguminosae), which includes several important forage legumes of temperate and subtropical regions, is characterized by small chromosomes of uniform size. Cytogenetic mapping of the two multigene families coding for 18S-5.8S-26S rRNA and 5S rRNA in white clover (T. repens)and seven closely related species/subspecies was carried out using fluorescencein situhybridization (FISH). The 18S-26S rDNA, generally confined to nucleolus organizer regions (NORs), was consistently located proximally on the shorter arm of one or two metacentric or submetacentric chromosome pairs. DAPI-negative, diffused and highly stretched 18S-26S FISH signals, representing previously transcriptionally active NORs, were observed up to late metaphase. The flanking condensed ends of the diffused 18S-26S rDNA signals were DAPI-positive and represented perinucleolar knobs which are presumed to be transcriptionally inactive. In interphase cells, the decondensed NOR chromatin passed through the nucleolar domain. In one species (T. isthmocarpum),in addition to two NOR chromosome pairs, 18S-26S signals were also located on a third pair, with no apparent NOR association. The 5S rDNA signals were located proximally on a long arm in six species and were syntenic to the NORs in all these species exceptT. ambiguumandT. nigrescensssp.petrisavii.In two species (T. hybridum, T. isthmocarpum),5S sequences occurred in the chromosomal satellites aligned distally to the NOR sequences. The relative distribution patterns of the two types of tandemly repeated DNA sequences were species-specific except forT. nigrescensssp.petrisaviiandT. ambiguum,both of which showed identical hybridization patterns. The two types of rDNA sequences have provided molecular markers for individual identification of a set of clover chromosomes. Present findings support the allotetraploid origin of white clover (T. repens)and suggest thatT. nigrescensssp.petrisaviimay be one of the present day diploid ancestors.

Physical mapping of 45S rRN A gene loci in the cucumber (Cucumis sativusL.) using fluorescencein situhybridization

Abstract The fluorescence in situ hybridization (FISH) using two different rDNA probes (18S and 45S) was applied to cucumber (Cucumis sativus L.) chromosomes. After counterstaining with 4’-6-diamidino-2-phenylindole (DAPI), all chromosomes showed visible bands similar to C-banding pattern and could be identified. The 18S and 45S rDNA signals appeared at the pericentromeric regions of chromosomes 1, 2 and 5. After silver staining to identify the nucleolar organizer region (NOR), the mitotic cells had two to four dots in nucleoli at interphase, two bands at the pericentromeric regions of two larger chromosomes at mid-metaphase. Although no NOR-bands appeared clearly at the prometaphase stage, certain silver-stained chromosomes were identified. DAPI staining made it possible to identify the silver-stained chromosomes at mid-metaphase and suggested that chromosome 1 or 2 had a silver-stained band at mid-metaphase.

Localization of 5S rDNA and 18S-5.8S-25S rDNA probes in &lt;i&gt;Crinum latifolium&lt;/i&gt; L. genome

Two conserved repeats viz. 5S rDNA and 18S-5.8S-25S rDNA were used as probes for physical mapping in the genome of Crinum latifolium L. (2n = 3X = 33). Nine out of 11 5S rDNA signals were found in three chromosomes of group VI at metaphase. The 5S rDNA gene array were not AT-rich. Four 18S-5.8S- 25S rDNA signals at interphase, prophase and metaphase showed their stable nature. The contraction and expansion of these signal regions proved the euchromatic nature of 18S-5.8S-25S rDNA. The 5S rDNA and the 18S-5.8S-25S rDNA signals provided useful cytogenetic markers for C. latifolium genome. Patterns of 5S rDNA signals suggested the possible structural aberration in the genome. Although the 5S rDNA sites suggested C. latifolium as autotriploid, 18S-5.8S-25S rDNA sites did not conclude anything about the nature of genome. Unequivocal determination of the nature of genome in C. latifolium remained unanswered. Key words: rDNA FISH; Crinum latifolium DOI: 10.3329/bjb.v39i2.7481 Bangladesh J. Bot. 39(2): 199-205, 2010 (December)