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Idiogram of chromosomes in the karyotypes of 13 B. napus varieties showing all possible positions of C-bands (black segments), 26S rDNA (green) and 5S rDNA (red). The chromosomes are grouped into metacentric, submetacentric and acrocentric types.
Source publication
The application of DNA intercalator 9-aminoacridine allowed us to increase the resolution of chromosome C-banding and DAPI-banding patterns and to investigate chromosomal polymorphism in karyotypes of seven spring and six winter rape varieties. It was shown that the pericentromeric and intercalary C-bands of most of the chromosomes in spring rape w...
Context in source publication
Context 1
... of DNA 9-AMA allowed us to obtain less condensed chromosomes containing more inter- calary C-bands (figure 1). Accordingly, we were able to identify chromosomes in karyotypes of the examined vari- eties, to investigate intraspecific C-band polymorphism and also to construct an idiogram of the rapeseed genome with account of polymorphism of chromosome C-banding pat- terns ( figure 3). Comparative analysis of rapeseed chromo- somes revealed C-band polymorphism in the examined vari- eties. ...
Citations
... Four varieties of winter and spring rapeseed (Podmoskovniy, Vikros, VIK 2 and Severyanin) were studied by Byelorussian researchers to identify the gene alleles determining the concentration of oleic and linolic acids in rapeseed oil (Lemesh et al., 2015). The same varieties were investigated to detect the DNA markers of the genes responsible for erucic-acid synthesis (Amosova et al., 2014). Microsatellite markers were used to study the genetic polymorphism of Russian varieties Ratnik and SNK-198 (Satina, 2010) as well as the genetic homogeneity of spring rapeseed varieties Bulat and Forward (Rogozhina et al., 2015). ...
Abstract. Rapeseed (Brassica napus L.) and turnip rape (B. rapa L. subsp. campestris (L.)) are important agricultural plants widely used for food, fodder and technical purposes and as green manure. Over the past decades, a large num-ber of perspective varieties that are being currently cultivated in every region of Russia have been developed. To increase the breeding eff iciency and facilitate the seed production, modern molecular-genetic techniques should be introduced as means to estimate species and varietal diversity. The objective of the presented research study was to investigate DNA polymorphism of the rapeseed and turnip rape varieties developed at Federal Williams Research Cen-ter of Forage Production and Agroecology and detect informative markers for varietal identif ication and genetic cer-tif ication. To genotype 18 gDNA samples, 42 and 25 combinations of respective SSR and SRAP primers were used. The results obtained demonstrate that SRAP markers were more effective for polymorphism analysis: 36 % of the tested markers revealed genetic polymorphism compared with only 16.7 % of microsatellite loci. Molecular mar kers to detect diffe rences at interspecif ic and intervarietal levels have also been found. For the investigated set, such microsatellite loci as Na12A02, Ni2C12, Ni02-D08a, Ra02-E01, Ni03H07а and SRAP-marker combinations as F13-R9, Me4- R7, F11-Em2, F10-R7, F9-Em2 and F9-R8 proved to be informative. Application of the two marker techniques made it possible to de-tect a higher level of DNA polymorphism in plants of different types (spring and winter varieties) if compared against the intervarietal differences within a species or a group. According to Nei’s genetic diversity index, in the cluster of winter rapeseed, VIK 2 and Gorizont varieties had the longest genetic distance, and in the spring cluster, these were Novosel and Veles. A high level of similarity was found between Vikros and Bizon winter rapeseed varieties. The results obtained have a high practical value for varietal specif ication of seed material and genetic certif ication of rapeseed and turnip rape varieties.
Key words: forage crops; Brassica napus L.; B. rapa L. campestris; bulk samples; genetic polymorphism; SSR mar kers; SRAP markers.
... and this species has simple and uniform C-banding patterns, that represented mostly by pericentromeric and telomeric HC bands. In another study Amosova et al. (2014) examined chromosomal and genetic polymorphism of 13 B. napus by studying C-banding and DAPI banding. A further declaration of relationships at chromosomal level and genetic control of meiosis in family Brassicaceae by utilizing GISH (Ali et al., 2004;Xiong & Pires, 2011). ...
... Knowledge of the co-segregation of alleles and their genetic linkages with the desired locus can be verified through the identification of loci using molecular marker systems. The number of loci, their positions, and their order within a chromosome was determined using various marker systems [(randomly amplified polymorphic DNA (RAPD), restriction fragment length polymorphism (RFLP), amplified fragment length polymorphism (AFLP), and simple sequence repeats (SSR), etc.)] (Nagaoka et al., 2010;Li et al., 2012;Amosova et al., 2014;Su et al., 2015). However, the efficacy of markers largely depends upon the number of amplified alleles and their comparative rate present on the genome. ...
Randomly amplified polymorphic DNA (RAPD) is a tremendously convenient approach used to discriminate between Brassica species owing to its accuracy and speed. RAPD primers generate adequate genetic information that can be used in the primer-marker system. In this work, twenty RAPD-PCR based markers were executed to generate polymorphic data, like polymorphic information content (PIC), mean resolving power (MRP), resolving power (RP), effective multiplex ratio (EMR), and marker index (MI) for the first time and genetic distance among and between six Brassica species were calculated. Our results indicated that 20 primers produced a total of 231 scored band and generated 87% polymorphic bands. Average PIC, MRP, RP, MI, and EMR values were 0.088, 0.65, 6.7, 0.78, and 8.9, respectively. PIC showed an overall negative correlation with MRP, RP, MI, and EMR, whereas MRP, RP, and EMR, were positively correlated with each other. Genetic identities ranged from 41.99% (between Brassica napus and Brassica oleracea) to 68.83% (between Brassica campestris and Brassica oleracea). Dendrogram results showed no clustering between species except between Brassica campestris and Brassica nigra. Nevertheless, these results will be helpful to acquire useful information about the markers and their use to determine the genomic structures of Brassica species. Further, based on genetic distance and polymorphic information, new hybrids can be developed for effective oilseed production.
... Показано также, что хромосомное распределение и число кластеров 45S и 5S рДНК, а также транскрипционная активность рибосомных генов у особей одного и того же вида растений может меняться под влиянием различных факторов окружающей среды [30][31][32][33]. У видов рода Brassica выявлен внутривидовой полиморфизм по хромосомной локализации кластеров 45S и 5S рДНК [34][35][36][37][38]. В кариотипах озимых сортов рапса обнаружена сортовая специфичность сочетаний полиморфных вариантов хромосом и особенности хромосомного распределения 45S и 5S рДНК [39,40]. ...
... Изменение числа локусов 45S рДНК в зависимости от мест произрастания популяций наблюдали в кариотипах и других видов растений, в том числе Phaseolus vulgaris [27] и видов рода Capsicum [29]. Внутривидовая изменчивость числа сайтов 45S и 5S рДНК обнаружена в кариотипах сортов озимого и ярового рапса, адаптированных к определенным климатическим зонам [39]. В результате проведенного нами исследования показано, что в кариотипах изученных масличных, листовых и кормовых озимых сортов B. rapa повышен уровень хромосомного полиморфизма по числу и распределению сайтов рибосомных генов. ...
Возделывание хозяйственно ценного вида Brassica rapa L. в регионах рискованного земледелия требует получения сортов, устойчивых к низким температурам. Механизм низкотемпературного стрессового ответа представляет собой сложную сеть процессов, которая затрагивает экспрессию многих
генов, включая рибосомные, а также коррелирует с хромосомной изменчивостью растений. В представленной работе методами FISH- и ПЦР-анализа впервые проведено изучение внутривидовой изменчивости хромосом по числу и локализации кластеров 45S и 5S рДНК генов, а также комплекса молекулярных маркеров холодоустойчивости у озимых сортов B. rapa, районированных в регионах рискованного земледелия. В результате исследования выявлены SSR-маркеры (Na10-CO3, BrgMS5339-1) и SCAR-маркеры (BoCCA1-F/BoCCA1-1R1, BoCCA1-F/BoCCA1-2R1), пригодные для диагностики устойчивых и восприимчивых к холоду генотипов B. rapa. У масличных и листовых сортов обнаружено большое число маркеров холодоустойчивости и высокий уровень
полиморфизма по распределению на хромосомах кластеров 45S и 5S рДНК, в том числе наличие гетероморфных вариантов хромосом. У кормовых сортов выявлены менее устойчивые к холоду генотипы и меньше полиморфных вариантов хромосом с сайтами рДНК. Таким образом, показано, что наименее устойчивые генотипы обладают меньшим хромосомным полиморфизмом по распределению кластеров рибосомных генов и наоборот. Полученные результаты могут представлять интерес для практической селекции при выведении новых, устойчивых к низким температурам сортов B. rapa.
... Due to small rapeseed chromosomes (1.53-3.30 μm) [43], the detailed chromosomal analysis is still problematic and needs special approaches, e.g., chromosome elongation with the use of DNA intercalators, application of chromosomal markers allowing identification of individual rapeseed chromosomes and their subgenomic affiliation [44][45][46][47]. Comprehensive study of the genotypic variability in mutant rapeseed lines in combination with the karyotype structure analysis (chromosomal complements in A and C subgenomes, the presence of chromosome rearrangements, chromosome substitutions and additions), description of phenotypic and biochemical variability was not performed. ...
... After the FISH procedure, chromosome slides were stained with 0.1 μg/mL DAPI (4 0 ,6-diamidino-2-phenylindole) (Serva, Heidelberg, Germany) in Vectashield mounting medium (Vector laboratories, Peterborough, UK). DAPI-banding analysis was used as an additional parameter for the identification of individual chromosomes [46,47]. ...
... In karyotypes, the cytological numerical designation of the chromosomes of A and C subgenomes was according to Levan's criterion [55]. Additionally, the identification of chromosomes and genome affiliation were performed based on the chromosome morphology, revealed chromosome markers as well as earlier described data [46,47,56,57]. The meiotic chromosome preparations were analyzed as described previously [51]. ...
The phenotypic, biochemical and genetic variability was studied in M2-M5 generations of ethyl methansulfonat (EMS, 0.2%) mutagenized rapeseed lines generated from canola, '00', B. napus cv. Vikros. EMS mutagenesis induced extensive diversity in morphological and agronomic traits among mutant progeny resulted in selection of EMS populations of B. napus- and B. rapa-morphotypes. The seeds of the obtained mutant lines were high-protein, low in oil and stabilized in contents of main fatty acids which make them useful for feed production. Despite the increased level of various meiotic abnormalities revealed in EMS populations, comparative karyotype analysis and FISH-based visualization of 45S and 5S rDNA indicated a high level of karyotypic stability in M2-M5 plants, and therefore, the obtained mutant lines could be useful in further rapeseed improvement. The revealed structural chromosomal reorganizations in karyotypes of several plants of B. rapa-type indicate that rapeseed breeding by chemical mutagenesis can result in cytogenetic instability in the mutant progeny, and therefore, it should include the karyotype examination. Our findings demonstrate that EMS at low concentrations has great potential in rapeseed improvement.
... The seeds were germinated in Petri dishes on the moist filter paper at room temperature. Root tips (of 0.5-1 cm) were excised and stored for 16-20 h in ice-cold water with 1 µg/mL of 9-aminoacridine (9-AMA) (Sigma, St. Louis, USA) to inhibit chromosome condensation process and accumulate prometaphase chromosomes 51,61,62 . After the pre-treatment, the root tips were fixed in ethanol:acetic acid (3:1) fixative for 48 h at room temperature. ...
The morphological, meiotic and chromosomal variability were studied in two cultivars of Calendula officinalis L. and their mutant lines obtained though chemical mutagenesis using diethyl sulphate (DES) (0.04%, 0.08%) and dimethyl sulphate (DMS) (0.025%, 0.05%). The studied cultivars displayed different sensitivity to DMS and DES mutagens. More M1 plants with morphological changes were observed in C. officinalis cv. ‘Zolotoe more’ than in cv. ‘Rajskij sad’. DMS and DES at low concentrations had positive effects on main agro-metrical traits in both cultivars including plant height, inflorescence diameter and number of inflorescences per plant. Dose-dependent increase in number of various meiotic abnormalities was revealed in both mutant lines. Comparative karyotype analysis and FISH-based visualization of 45S and 5S rDNA indicated a high level of karyotype stability in M1 and M2 plants. Seed treatments with DMS and DES at certain concentrations resulted in higher yields of inflorescences in M1 plants compared to the control. In M2 generation, dose-dependent reduction in the yields of inflorescences was observed. Our findings demonstrate that DMS and DES at low concentrations have great potential in calendula mutation breeding.
... Due to high copy number, detection of rDNAs is highly reproducible and provides valuable information concerning chromosomal evolution [34]. In plant genomes, the copy numbers and chromosomal distribution of rDNAs can vary rapidly even within intraspecific taxa and can therefore provide chromosomal landmarks for genome plasticity [34][35][36]. ...
The ontogenesis and reproduction of plants cultivated aboard a spacecraft occur inside the unique closed ecological system wherein plants are subjected to serious abiotic stresses. For the first time, a comparative molecular cytogenetic analysis of Pisum sativum L. ( Fabaceae ) grown on board the RS ISS during the Expedition-14 and Expedition-16 and also plants of their succeeding (F1 and F2) generations cultivated on Earth was performed in order to reveal possible structural chromosome changes in the pea genome. The karyotypes of these plants were studied by multicolour fluorescence in situ hybridization (FISH) with five different repeated DNA sequences (45S rDNA, 5S rDNA, PisTR-B/1, microsatellite motifs (AG) 12 , and (GAA) 9 ) as probes. A chromosome aberration was revealed in one F1 plant. Significant changes in distribution of the examined repeated DNAs in karyotypes of the “space grown” pea plants as well as in F1 and F2 plants cultivated on Earth were not observed if compared with control plants. Additional oligo-(GAA) 9 sites were detected on chromosomes 6 and 7 in karyotypes of F1 and F2 plants. The detected changes might be related to intraspecific genomic polymorphism or plant cell adaptive responses to spaceflight-related stress factors. Our findings suggest that, despite gradual total trace contamination of the atmosphere on board the ISS associated with the extension of the space station operating life, exposure to the space environment did not induce serious chromosome reorganizations in genomes of the “space grown” pea plants and generations of these plants cultivated on Earth.
... In FISH assays, mitotic chromosome spreads were prepared from plant root meristem according to the technique developed previously for plant species with small chromosomes Amosova et al. 2014). For meiotic chromosome preparation, young floral buds were fixed in Carnoy's solution for 30 min at 4°C and then chromosome spreads were prepared as previously described . ...
... A M-I: (n = 10 II ? 1 IV ); B A-I: normal distribution of chromosomes in the cell (14:14); C, D A-I: chromosome lagging; E A-I: non-uniform chromosome distribution within the cell; F A-II: nonuniform chromosome distribution within a cell; G A-II: chromosome lagging; H tetrad; I hexadprocess of chromosome condensation, allowed us to accumulate prometaphase chromosomes and obtain longer chromosomes in the spreads (1.5-3 lm) resulting in rather informative chromosome DAPI-banding patterns. In P. alba karyotypes, we observed large pericentromeric and also small polymorphic telomere and intercalary DAPI bands, and such patterns are typical for plant species having small chromosomes(Guerra 2000;Pinto-Maglio 2006;Samatadze et al. 2012;Yurkevich et al. 2013;Amosova et al. 2014). ...
Potentilla alba L. is a valuable medicinal plant widely used in folk and traditional medicine and particularly promising in complex treatment of thyroid pathology. Natural resources of this species are insufficient due to ever-growing use in contemporary medicine. Comprehensive investigations of different P. alba populations are essential for the successful extension of P. alba plantings. Aiming for a better understanding of karyotype structure, chromosome behaviour in meiosis and developing new diagnostic characters, we performed molecular cytogenetic characterization and leaf structure and ultrastructure analyses of two introduced P. alba samples originating from different habitats. Based on chromosome morphology, distribution of 45S/5S rDNA and DAPI-banding patterns, all chromosomes in the karyotypes were identified and the P. alba chromosomal idiogram was constructed. Our findings confirmed P. alba karyotype stability and also revealed several diagnostic characters of this species: the features of cells of upper and lower leaf epidermis, the presence of calcium oxalate druses and three types of leaf indumentum, essential for evaluation of genetic diversity in different populations, validation of raw materials and further selection progress. The meiotic abnormalities were detected probably related to low pollen activity and indicated the advantages of vegetative propagation in the development of a P. alba plantation system.
... Since B. napus chromosomes are small in size (1.53-3.30 lm) (Hasterok and Maluszynska 2000), their investigation is still problematic and needs special approaches such as chromosome elongation by using DNA intercalators as well as application of chromosomal markers which could allow the identification of individual rapeseed chromosomes and also their subgenomic affiliation (Howell et al. 2008;Xiong and Pires 2011;Amosova et al. 2014;Zemtsova et al. 2014). ...
... In the present study, we observed segregation of morphological traits among S2 hybrids though they were karyotypically normal (2n = 38). The performed molecular cytogenetic analysis showed that chromosomal localization of 45S and 5S rDNA loci revealed in S2 hybrids corresponded to that found in natural rapeseed Zemtsova et al. 2014) as well as in genomes of the parental species despite the polymorphic nature of rDNA loci in B. napus (Hasterok et al. 2001;Ali et al. 2005;Hasterok et al. 2006;Amosova et al. 2014). The phenotypic changes in S2 hybrids have probably resulted from the alterations in regulation of gene expression in the synthetic rapeseed genome. ...
Resynthesized Brassica napus L. is an important source for broadening genetic diversity and producing lines with desired characteristics. It is also a fine model to study the processes of genomic reorganizations in recently formed polyploids. We firstly performed molecular cytogenetic characterization of newly resynthesized rapeseed (B. rapa ssp. narinosa × B. oleracea ssp. capitata) and its parental species, and also examined genomic changes in hybrids of the succeeding generations grown under pressure of selection of yellow-seeded progeny. For karyotype studies, FISH/GISH with 45S, 5S rDNA, C genome specific BoB014O06 BAC clone and genomic DNA of parental B. rapa was performed. Synthetic S0–S2 hybrids had common rapeseed karyotypes (2n = 38) including 14 loci of 45S rDNA sites and 10 loci of 5S rDNA. Progeny selection led to gradual deletion of C genome chromosomes in hybrid karyotypes. So, in karyotypes of S6 and S7 hybrids, the chromosome number was reduced to 2n = 20–22, and only chromosomes of A genome bearing 10–13 loci of 45S rDNA and 8–10 loci of 5S rDNA, variations in chromosome number, chromosome rearrangements as well as examples of trisomy and monosomy were revealed. Our findings indicate an enhanced genome instability in resynthesized rapeseed lines developed under the pressure of selection which might lead to chromosome rearrangements or/and deletions and even elimination of the whole parental genome in hybrids in succeeding generations. The approach can be useful for the development of rapeseed lines with trisomy, chromosome addition/substitution lines important for genetics and plant breeding.
... Brassica rapa, the presumed progenitor species of B. napus, has five loci (two major and three minor) per haploid set while the second presumed genome donor, B. oleracea, harbours two loci (Maluszynska and Heslop-Harrison, 1993). At the cytogenetic level, the cultivars of B. napus show genotypic differences in number, distribution and morphology of rDNA chromosomal loci (Snowdon et al., 1997;Fukui et al., 1998;Hasterok et al., 2001Hasterok et al., , 2006Ali et al., 2005;Amosova et al., 2014). Previous Southern blot hybridization revealed restriction fragments corresponding to both parents indicating Mendelian inheritance of rDNA in B. napus (Bennett and Smith, 1991;Waters and Schaal, 1996). ...
... Third, the C-genome rDNA sites were not totally blocked by the B. oleracea-specific IGS probe (Howell et al., 2008). Finally, chromosome banding showed interpopulation variation in the amount and distribution of heterochromatin adjacent to or overlapping with NORs (Amosova et al., 2014). ...
Background and aims
Brassica napus (AACC, 2n = 38, oilseed rape) is a relatively recent allotetraploid species derived from the putative progenitor diploid species Brassica rapa (AA, 2n = 20) and Brassica oleracea (CC, 2n = 18). To determine the influence of intensive breeding conditions on the evolution of its genome, we analysed structure and copy number of rDNA in 21 cultivars of B. napus, representative of genetic diversity.
Methods
We used next-generation sequencing genomic approaches, Southern blot hybridization, expression analysis and fluorescence in situ hybridization (FISH). Subgenome-specific sequences derived from rDNA intergenic spacers (IGS) were used as probes for identification of loci composition on chromosomes.
Key Results
Most B. napus cultivars (18/21, 86 %) had more A-genome than C-genome rDNA copies. Three cultivars analysed by FISH (‘Darmor’, ‘Yudal’ and ‘Asparagus kale’) harboured the same number (12 per diploid set) of loci. In B. napus ‘Darmor’, the A-genome-specific rDNA probe hybridized to all 12 rDNA loci (eight on the A-genome and four on the C-genome) while the C-genome-specific probe showed weak signals on the C-genome loci only. Deep sequencing revealed high homogeneity of arrays suggesting that the C-genome genes were largely overwritten by the A-genome variants in B. napus ‘Darmor’. In contrast, B. napus ‘Yudal’ showed a lack of gene conversion evidenced by additive inheritance of progenitor rDNA variants and highly localized hybridization signals of subgenome-specific probes on chromosomes. Brassica napus ‘Asparagus kale’ showed an intermediate pattern to ‘Darmor’ and ‘Yudal’. At the expression level, most cultivars (95 %) exhibited stable A-genome nucleolar dominance while one cultivar (‘Norin 9’) showed co-dominance.
Conclusions
The B. napus cultivars differ in the degree and direction of rDNA homogenization. The prevalent direction of gene conversion (towards the A-genome) correlates with the direction of expression dominance indicating that gene activity may be needed for interlocus gene conversion.
