Fluorescent in situ hybridization (FISH) has been a powerful technique to reveal chromosomal loci and distribution of DNA sequences in metaphase chromosomes. Three-dimensional FISH (3D-FISH) has also revealed the chromosome territory of interphase chromosomes in the nucleus. Recently, 3D-FISH has played a supportive or complementary role to confirm the genome organization and topologically associated domains, which are provided by chromosome conformation capture techniques.
Using a base-specific staining method, three Japanese cultivars of Momordica charantia were investigated. The present method using enzymatic digestion produced a high level of well-spread metaphase chromosome complements. The cultivars investigated showed the same somatic chromosome number 2n=22. In condensation behavior from prophase to metaphase, most chromosomes of all cultivars in this study had early condensing segments at the proximal regions. Moreover, at prophase or earlier stage of metaphase, most chromosomes observed had decondenced segments at the distal regions in both arms. Judging by the determination of centromere positions, the chromosome complements at mitotic metaphases in three cultivars consisted of 22 metacentric chromosomes. Moreover, similar sizes and shapes of the chromosomes indicated that the karyotypes of M. charantia cultivars were symmetrical. In contrast, fluorescent staining showed different sizes and numbers of chromycin A3 (CMA) positive and 4′,6-diamidino-2-phenylindole (DAPI) negative (CMA+DAPI−) satellites among cultivars. Four CMA+DAPI− satellites were observed in M. charantia ‘Abashi-goya’ and M. charantia ‘Naga-goya,’ while two satellites were obseved in M. charantia ‘Shiro-goya.’ All satellites were located at one end of sat-chromosomes. In ‘Shiro-goya,’ a large size difference of satellites was found between two sat-chromosomes, which might be homologous chromosomes with polymorphism. Moreover, faintly CMA+DAPI− sites were shown at the proximal regions or primary constrictions of most chromosomes in all cultivars.
The present study is a part of our research project on the meiotic studies of some Monocots from the Rajasthan area. Rajasthan, the hot desert of India, has low biodiversity. The present paper deals only with the 19 species of the tribe Paniceae out of these 3 species, viz., B. mutica (n=14), Eriochloa procera (n=27) and Panicum typheron (n=9), are new euploid chromosome reports (two for the world and one for India). Out of these, abnormal meiosis is found in Brachiaria reptans (n=7), B. mutica (n=14), Eriochloa procera (n=27) and Paspalum paspalodes (n=30), whereas meiotic course is normal in the others.
The present article has reported a detailed karyotype analysis and 4C nuclear DNA estimation in different cultivars and populations of Clitoria ternatea L. of Fabaceae. This plant, widely distributed in tropical regions in India, is a climber and is an economically important medicinal plant. The karyotype study has revealed a constancy in the diploid somatic chromosome number (2n=16) in all these cultivars and populations. However, the nature and number of nucleolar chromosomes varied. The centromeric chromosomes were either metacentric or sub-metacentric in all these cultivars. The total chromosome length and volume differed among these cultivars and populations having the same somatic chromosome number. The differential condensation of chromosomes and the association of variable amounts of proteins in chromosome composition might have been responsible for such differences. In situ cytophotometric studies revealed an intermediate genome size in this species. This is the first report on the detailed karyotype and genome size of cultivars of Clitoria ternatea L.
The present study pertains to the cytomorphology of 19 species belonging to nine families of the subclass Gamopetalae from Hamirpur district (H. P.). Out of these, 5 species are new or varied chromosomes reports. Campanula benthamii (2n=2x=28) and Leucas hyssopifolia (2n=2x=22) are the first ever chromosome counts for the species. Spilanthes acmella (2n=6x=78) and Borreria articularis (2n=4x=52) are varied cytotypes. Meanwhile, the B chromosome is reported for the first time in Leucas lanata (2n=22+0-1B). Almost all of the accessions (with disparity in percentage) show laggards, chromosome stickiness and cytomixis from early prophase to telophase-II, leading to the formation of aneuploid cells or meiocytes with double the normal chromosome number. Such meiotic abnormalities produce unreduced pollen grains and reduce pollen viability.
Stichococcus Nägeli is a unicellular green algal species that belongs to the Trebouxiophyceae. Stichococcus bacillaris Nägeli NIES-3639 propagates by binary fission, but can form a filament without separating into two daughter cells. Transmission electron microscopy showed that cytokinesis starts according to invagination of the plasma membrane at the plane of division. Synthesis of the daughter cell wall occurs on the surface of invaginating plasma membrane during the protoplast division phase. The mother cell wall may cleave after protoplast division in either the single-cell or filament state. Field emission scanning electron microscopy revealed scars of cleaved mother cell walls from the single-cell state, two-cell state after cell division, and four-cell state during filament formation. Because the mother cell wall was cleaved, the delay of the daughter cell separation may have caused filament formation. Since this cell division cycle repeats rapidly, it may begin before the separation of the daughter cells yielded by the former cell cycle.
The present study was conducted to understand the effects of different doses of gamma radiations on the cytological behavior of common wheat (Triticum aestivum var. WH-147) in M1 generation. Progenies were raised from the seeds of the selected cultivar treated with six different doses of gamma radiations, viz., 5, 10, 15, 20, 25 and 30 kR at Bhabha Atomic Research Center, Mumbai. Cytological studies were carried out in the treated as well as in control materials. The radiations have shown a significant effect causing different types of chromosomal anomalies such as stickiness, univalents, laggards, fragments, bridges, precocious segregation, and micronuclei. In general, an increasing spectrum of aberrant cells was observed along with the increasing intensity of gamma radiations. The cytological analysis also revealed that the most frequent anomaly dominated was chromosomal stickiness followed by other irregularities. In addition, 25 and 30 kR doses of gamma radiations were the most potent doses, inducing higher frequency of cytological aberrations in pollen mother cells of wheat, suggesting that gamma rays could be successfully employed for creating additional genetic variability in the natural gene pool of common wheat.
In Saccharomycodes ludwigii, two spores with opposite mating types are tethered together by a ledge at the poles of a lemon-shaped apiculate cell. In the present study, we performed sporulation culture of spheroplasts of S. ludwigii in order to investigate how cell shape affects the formation of spore pairs. Each spore pair in whole cells or in spheroplasts was isolated using a micromanipulator, and the colonies derived from the germination of spore pairs were tested for sporulation ability and mating type. In terms of the results, among 55 colonies derived from spore pairs in whole cells, only one colony was haploid and the other 54 colonies were diploid. In contrast, among 55 colonies derived from spores in spheroplasts, 10 colonies were haploid and the other 45 colonies were diploid. These results, together with microscopic observations, suggest that the lemon-shaped cell morphology is important for the formation of normal spore pairs.
In the present paper, karyotype analyses are reported for 12 taxa of Matthiola from Turkey. Matthiola trojana Dirmenci, Satıl & Tümen, M. odoratissima (Pall.) R.Br., M. anchonifolia Hub.-Mor., M. montana Boiss., M. fruticulosa (L.) Maire ssp. fruticulosa and M. ovatifolia (Boiss.) Boiss. counted 2n=12 chromosomes, and M. incana (L.) R.Br., M. longipetala (Vent.) DC. ssp. bicornis (Sibth. et Smith) P. W. Ball, M. longipetala (Vent.) DC. ssp. longipetala, M. longipetala (Vent.) DC. ssp. pumilio (Sibth. & Smith) P. W. Ball, M. sinuata (L.) R.Br., and M. tricuspidata (L.) R.Br. showed 2n=14 chromosomes. The idiograms and karyomorphometric data obtained by using Image Analysis System (Bs200Pro). The chromosome number of M. montana was determined for the first time. Also, karyotype asymmetry index such as MCA, CVCL and CVCL were measured for the reconstruction of karyological relationships.
The six Abies species, A. laciocarpa, A. veitchii, A. sachalinensis, A. mariesii, A. faxoniana and A. georgei, were investigated for their chromosomes by the fluorescent banding method using fluorochromes chromomycin A3 (CMA) and 4′,6-diamidino-2-phenylindole (DAPI). All six species have 2n=24 chromosomes and a similar karyotype consisting of seven pairs of long metacentric chromosomes and five pairs of shorter submeta- and subtelocentric chromosomes, supporting previous studies. Eight clear CMA-bands appeared at the interstitial region of one arm of long metacentric chromosomes in all species, and in A. veitchii and A. sachalinensis, their number varied from six to eight among plants and/or populations. A weak CMA-band appeared at the interstitial region of one chromosome pair, while a weak CMA-band appeared at the proximal region in some species. In most species DAPI did not produce clear bands, and sites of clear CMA-bands were DAPI negative. Only A. mariesii showed many DAPI-bands at the interstitial and/or centromeric regions of several chromosomes. A few weak DAPI-bands appeared in some other species. The fluorescent banding and FISH patterns reported in Abies species were compared and discussed with taxonomic treatment and molecular phylogeny of Abies.
Eukaryotic cells contain three types of nuclei: the cell nucleus, the mitochondrial nuclei (nucleoids), and plastid nuclei (nucleoids). Here, we report that the chlorophyte strain Medakamo hakoo 311 has the smallest cell nucleus genome among the free-living eukaryotes analyzed to date. The size of the nuclear genome was analyzed by staining with 4′,6-diamidino-2-phenylindole (DAPI), followed by fluorimetry using a video-intensified microscope photon-counting system (VIMPCS) and fluorescence microscopy. The exact genome size of M. hakoo remains elusive because of the low permeability of the thick algal cell wall to DAPI. To improve the DAPI staining method for M. hakoo, we added a microwave oven heating step. We examined the genome sizes of M. hakoo, Cyanidioschyzon merolae, Saccharomyces cerevisiae (n, 2n), and Chlorella variabilis 1 N by DAPI staining, which included a microwave oven heating step, followed by fluorimetry with the VIMPCS. The new method of DAPI staining that included a microwave oven heating step improved the staining efficiency of C. variabilis and M. hakoo nuclei, and slightly improved the staining efficiency of S. cerevisiae and C. merolae nuclei. Using well-stained samples and the cell nucleus genome size of C. merolae (16.5 Mb) as the standard, the cell nucleus genome sizes of M. hakoo, S. cerevisiae (n), and C. variabilis were determined as 9.2, 20.2, and 35.9 Mb, respectively. The results are consistent with previous reports that M. hakoo cell has the smallest genome among the free-living eukaryotes evaluated to date.
The present research has been carried out on Adiantum capillus-veneris L., a well renowned medicinal fern, to study its detailed meiotic behaviour in 14 populations collected from different parts of Northwest India. All the populations uniformly revealed the presence of 30 bivalents at M-I as reported by previous workers. However, irregular meiotic behaviour characterized by the presence of stickiness, unoriented bivalents, laggards, bridges, interbivalent connections, secondary associations, cytomixis, formation of micronuclei leading to abnormal microsporogenesis, i.e., spore sterility, were investigated. The abnormal meiotic course in some studied populations has a significant effect on spore viability, i.e., reduced spore fertility. To our knowledge, this is the first ever population based report on abnormal meiotic behaviour of this species.
Detailed meiotic investigations were performed in three accessions of Meconopsis aculeata scored from high altitudinal regions in the Northwest Himalayas. These accessions exhibit the same chromosome number of n=28 and exist at tetraploid level. We report here the occurrence of chromatin stickiness and abnormal spindle activity in the presently studied accessions. Consequent to chromatin stickiness and abnormal spindle activity, various irregularities were observed at different stages of meiosis. These include inter-bivalent connections, out-of-plate bivalents, laggards and chromatin bridges during anaphases/telophases resulting in the formation of abnormal sporads like triads, tetrads with micronuclei and polyads. The products of such sporads yielded heterogeneous-sized fertile and sterile pollen grains. Chromatin stickiness and abnormal spindle activity causing meiotic irregularities, abnormal sporads and pollen malformation seem to be the result of low temperature stress conditions prevailing in the area when the plants enter the flowering stage.
Diploid XP cells were exposed to demecolcine for three days and then released from this exposure. Tetraploid XP cells were successfully established 60 d after drug addition. The doubling time of the tetraploid cells was 18 h, identical to that of diploid cells. The tetraploid cells had 92 chromosomes and double the DNA content of the parent diploid cells. The cell volume of tetraploid cells was about twice that of the diploid cells. The duration of the G1, S and G2/M phases was almost the same as for diploid cells. It was concluded that the established tetraploid XP cells have cellular characteristics similar to those of diploid XP cells. The rate of survival after UV irradiation was lower in tetraploid XP cells than in diploid XP cells.
Lotus is a leguminous and cosmopolitan genus in the Loteae tribe consisting of more than 200 species. The number of chromosomes has been reported for many Lotus species; however, molecular studies have focused only on a few important species of this genus. The present study was conducted to estimate the genome size and ploidy levels of 28 Lotus accessions, and to identify their genetic diversity using inter-simple sequence repeat (ISSR) analysis. The chromosome number of 16 accessions agreed with previous reports (except for Lotus salsuginosus), while that in 11 accessions were reported here for the first time. The smallest nuclear DNA content was identified in the diploid, Lotus unifoliolatus (0.28±0.01 pg C−1). In contrast, the tetraploid, Lotus australis, had a genome size of 1.28±0.03 pg C−1, representing a five-fold difference in genome size among the Lotus species. When expressed as a per Cx value, Lotus species genome sizes ranged from 0.28 pg Cx−1 in L. unifoliolatus to 0.71 pg Cx−1 in Lotus wrightii, representing a 2.5-fold difference. There was no relationship between genome size and chromosome number or ploidy level; thus, genome size is species specific in the Lotus species. On ISSR analysis, a total of 379 fragments were generated with 12 primers, and all accessions were grouped into four clusters by phylogenetic analysis. The results of this investigation will be useful for plant breeders attempting to expand the genetic variation found in this species by crossbreeding using these resources.
The present study was performed to evaluate the role of Aloe vera L. leaf extract (AvLE) on some physiological and cytogenetical parameters of Allium cepa L. seeds exposed to salinity. The radicle length of the seeds germinated in the medium with AvLE alone increased as compared with ones of the control seeds germinated in distilled water medium, while their radicle number and fresh weight reduced according to the control. In addition, the germination percentage of the mentioned seeds statistically was the same as the control seeds. Furthermore, the seeds germinated in the media containing 0.1 mg/L AvLE alone showed a significant increase of mitotic index, although they exhibited a higher number of chromosomal aberrations and micronucleus (MN) formation as compared with the seeds germinated in control conditions. On the other hand, salt stress considerably inhibited the seed germination and seedling growth of A. cepa. Moreover, salinity markedly decreased the mitotic index in root tip meristems of the seeds and increased the number of chromosomal aberrations. The negative effect of salt on the seed germination, seedling growth, mitotic activity and MN formation was alleviated in dramatically varying degrees by AvLE application. In contrast, the detrimental effects of salt on the chromosomal aberrations greatly increased with this treatment. Consequently, we reported that the application of AvLE alone induced chromosomal aberrations and MN formation, which are an indicator of genotoxicity, and had cytotoxic activity in normal conditions. However, the frequency of MN is greatly reduced in root tip cells of the seeds germinated in the medium containing 0.1 mg/L AvLE +0.15 M NaCl.
Eleven varieties of Gossypium hirsutum L. (cotton) released by the Bangladesh Cotton Development Board, viz., CB-1 (Cotton Board-1), CB-2, CB-3, CB-4, CB-5, CB-6, CB-7, CB-8, CB-9, CB-10 and CB-11 were investigated cytogenetically by differential fluorescent banding for authentic characterization. The 11 cotton varieties represented a broad spectrum of variation for several phenotypic and agronomic traits. The 11 cotton varieties were found to possess 2n=52 chromosomes, of which most were metacentric. A wide range of CMA-positive bands (5–20) was found in the metaphase chromosomes of the 11 varieties. Different numbers of satellites, such as 2 (CB-4, CB-7), 3 (CB-6), 4 (CB-1) and 6 (CB-2, CB-3, CB-8, CB-10), were found after CMA-staining. The differential staining property of satellites revealed the stain-specific nature of these satellited portions. Entirely DAPI-fluoresced chromosomes were frequent in these varieties. The number, location and distributions of GC- and AT-rich repeats are specific for each variety. Some CMA- and DAPI-bands were so unique that these chromosomes could easily be isolated from the rest and may be used as marker chromosomes for the respective varieties. Fluorescent banding revealed the occurrence of genomic alteration within these varieties. Therefore, each variety could be characterized authentically by fluorescent banding analysis.
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