CYTOLOGIA
Online ISSN : 1348-7019
Print ISSN : 0011-4545
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Technical Note
Focus
  • Bingyi Liang, Hikaru Sato, Sachihiro Matsunaga
    2023 Volume 88 Issue 4 Pages 283-288
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Gene editing technology based on the clustered regularly interspaced short palindromic repeats (CRISPR) system has garnered widespread use in plant genomes because of its cost-effectiveness, efficiency, and simplicity. To avoid the integration of foreign genes and any DNA fragments into target cell genomes, researchers have developed a system that introduces in vitro-assembled ribonucleoproteins (RNPs) consisting of guide RNA (gRNA) and Cas protein into target cells, enabling direct genome editing. This system was designed to deliver RNPs through four distinct methods: polyethylene glycol (PEG)-mediated cell transfection, particle bombardment, electroporation, and lipid transfection. In recent years, CRISPR technology has been extensively applied for the genetic modification of plants, providing a strategic response to environmental challenges. Researchers have successfully established RNP genome editing systems in various plant species. Despite some remaining issues, the RNP genome editing system still shows significant promise for future applications in the production of non-genetically modified (non-GM) crops.

Regular Article
  • Murat Kürşat, Osman Gedik, Irfan Emre
    2023 Volume 88 Issue 4 Pages 289-294
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    The purpose of this work is to identify chromosomal numbers and various karyological characteristics of eight Euphorbia taxa grown in Turkey (E. heteradena Jaub. & Spach, E. altissima Boiss. var. altissima, E. macroclada Boiss., E. orientalis L., E. cheiradenia Boiss. & Hohen., E. seguieriana Neck. subsp. sequieriana, E. aleppica L., and E. chamaesyce L.). It was discovered that the Euphorbia taxa’s basic chromosome numbers were x=8, 9, 10, and 11. This work determined the Stebbins classification, ploidy levels, karyotype formula, chromosomal length range, total karyotype length, various karyotype asymmetry values, and the A1 and A2 in each taxon. In addition, the dispersion diagram was created using A1 and A2. Euphorbia taxa were revealed 2n=16 or 18 chromosomes apart from E. aleppica has 2n=20 and, E.chamaesyce has 2n=44, is polyploid. Also, they have median and submedian centromeric chromosomes. According to Stebbins classification, E. heteradena is 3A, E. altissima var. altissima, and E. orientalis are 2B, and the remaining species are 2A.

  • Chiaki Yaji, Riyo Kawaguchi, Hiroki Yasuhara
    2023 Volume 88 Issue 4 Pages 313-319
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Caffeine has long been known to be a specific inhibitor of plant cytokinesis. Caffeine inhibits callose deposition in the cell-plate, albeit via an unknown mechanism. We previously reported that the caffeine-sensitive processes that are required for callose deposition in the cell-plate are initiated at or before the onset of mitosis in cell-cycle-synchronized tobacco BY-2 cells. Because the content of UDP-glucose, which is a substrate for callose synthesis, reportedly increases prior to mitosis, we hypothesized that caffeine affects the generation and/or accumulation of UDP-glucose. In the present study, as a first step to verify this hypothesis, we examined whether caffeine inhibits cell-wall regeneration in protoplasts, which requires UDP-glucose. As a result, we found that caffeine inhibited the cell-wall regeneration of protoplasts in a dose-dependent manner, and that the effect was reversible. These results support the notion that the sites of action of caffeine are the generation and/or accumulation of UDP-glucose, rather than the previously proposed regulatory mechanisms mediated by Ca2+.

  • Tadashi Maruyama, Masashi Yamaguchi, Akihiro Tame, Takashi Toyofuku, H ...
    2023 Volume 88 Issue 4 Pages 321-329
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Supplementary material

    Dinoflagellates, including species of Tripos (formerly Ceratium) and Akashiwo, have two dissimilar flagella, a transverse flagellum, and a longitudinal one (LF). The LF of Tripos has been shown to perform two different motions, undulation, and retraction. The latter unique flagellar motion has been reported only from Tripos. In the present paper, we found for the first time that the LF of an unarmored dinoflagellate, Akashiwo sanguinea, also showed this unique motion. While the LF showed undulatory motion during swimming, physical stimulation of the cell induced its rapid retraction into the ventral groove (sulcus). After retracting, the LF re-extended and resumed undulation. Scanning electron microscopy of the retracted LF showed that it was folded at 3–4 µm intervals. Transmission electron microscopy showed that the flagellum has a complex structure, which consisted of a typical 9+2 microtubule-dynein system, a paraxonemal rod, and a fiber, which is similar to the contractile R-fiber of Tripos. A non-detergent extraction model of Akashiwo cells using EDTA-containing medium without a detergent showed that 1 mM Ca2+ induced the retraction of the LF, and that this activity was counteracted by Mg2+ at concentrations higher than 10 mM. The present data indicate that the LF of Akashiwo performs two dissimilar motions: undulation and retraction; that it contains two different motility systems: the 9+2 microtubule-dynein axoneme and the R-fiber-like fiber; and that Ca2+ plays an important role in the LF retraction.

  • Takahiro Katogi, Yuki Yoshida, Kaito Nakayama, Yoshikazu Hoshi, Shinic ...
    2023 Volume 88 Issue 4 Pages 339-346
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Supplementary material

    The genome size and karyomorphology of six accessions of Limosella aquatica L. in Japan were investigated. All six accessions had a chromosome number of 2n=40. The genome size of L. aquatica was estimated using flow cytometry (FCM) and 2C of the tetraploid genome size ranged from 1.38 pg in accession Shiranuka to 1.42 pg in accession Taiki (1,178–1,216 Mbp), with 1Cx estimated to be ca. 0.35 pg (300 Mbp). The GC contents of all accessions were less than 40%. The centromeres of all chromosomes were located at median positions, while secondary constrictions were found at the distal regions of two chromosomes, resulting in small chromatin segments as satellites. The chromosome complement showed a monomodal karyotype with a gradual decrease in the chromosome length, ranging from 1.3 to 0.6 µm. The average chromosome lengths of accessions in Hokkaido and Honshu were 0.9 and 1.0 µm, respectively. The accessions in Kumamoto were exceptions, showing chromosome lengths ranging from 1.6 to 0.8 µm. The DNA base-specific banding revealed that the pericentromeric regions of all chromosomes and the satellites were chromomycin A3 positive and 4′,6-diamidino-2-phenylindole negative (CMA+DAPI). The 18S rDNA fluorescence in situ hybridization (FISH) result also showed that the two signals were located at exactly the same positions as the CMA+DAPI satellites.

  • Yuki Tominaga, Takahiro Katogi, Sayuri Funakoshi, Ayano Mimura, Satomi ...
    2023 Volume 88 Issue 4 Pages 353-363
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Supplementary material

    Karyomorphological evaluations of the amphidiploid species Drosera tokaiensis (2n=60) and its parent species D. rotundifolia (2n=20) and D. spatulate (2n=40) were followed by an investigation of monochromatic light responses among these three species. The biseriate glandular trichome (BGT) maintained the characteristic cell morphology of each species, regardless of the differences in monochromatic light. The plant morphologies of all three Drosera species showed that petiole and stem growths were stimulated by blue light and inhibited by red light. In contrast, inflorescences of D. spatulata formed regardless of light conditions, but not D. rotundifolia. All Drosera species showed increased fresh weights when cultured under blue light, although the dry weight was not significantly different from that under red or white light, except for that of D. rotundifolia. The results of seven phenolic compounds of the three Drosera species revealed that milicitrin and quercimelin, lacking in D. rotundifolia, and ellagic acid low-detecting in D. spatulata, were present in D. tokaiensis. The contents of myricitrin and quercimelin in the Drosera extracts were higher under red light than under blue light for D. spatulata and D. tokaiensis. In contrast, the content of ellagic acid in the extracts was slightly higher under red light conditions than under blue light for D. tokaiensis. However, it was higher under blue light than under red light in D. rotundifolia, and other phenolic compounds also increased under blue light.

  • Yoshiko Kono, Ching-I Peng, Kazuo Oginuma, Hsun-An Yang, Kuo-Fang Chun ...
    2023 Volume 88 Issue 4 Pages 365-371
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    The previous chromosome information for 2n=24 and 2n=26 is limited to nine species in the Asian Begonia sections Diploclinium and Platycentrum. In this study, we report the first-ever chromosome counts for two species with 2n=24 and new chromosome counts for two species with 2n=26. Consequently, the total number of species with 2n=24 and 2n=26 has now increased to 13 species within the two sections, Diploclinium and Platycentrum, out of the 20 sections of Asian Begonia. We also provide the first karyotype report of these two species with 2n=24 and 2n=26 in sect. Platycentrum. Based on our findings and previous chromosome reports, which encompass 13 species with 2n=24 and 2n=26, we will discuss the chromosome evolution of 2n=24 and 2n=26 within sect. Platycentrum.

  • Woo-Cheol Kim, Hye-Rin Kim, Se-Mun Ahn, Kweon Heo
    2023 Volume 88 Issue 4 Pages 379-385
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Due to reproductive isolation, such as self-incompatibility, heterostyly, and higher genetic diversity within populations, the origin and evolution of buckwheat are difficult to study even though there are several cytological studies have been performed. In this study, Fagopyrum esculentum, F. esculentum ssp. ancestrale, F. tataricum, and F. tataricum ssp. potanini were used for karyotype analysis by double-color fluorescence in situ hybridization (FISH) using 5S and 45S rDNA probes. The karyotype analysis revealed that F. esculentum and F. esculentum ssp. ancestrale have the same karyotype formula, 6m+2sm (satellite, SAT). Conversely, F. tataricum and F. tataricum ssp. potanini possessed differing karyotypes, 6m+2sm (SAT) and 6m+1sm+1sm (SAT), respectively. From the FISH analysis, F. esculentum and F. esculentum ssp. ancestrale represented three pairs of 5S and 45S rDNA signals. F. tataricum and F. tataricum ssp. potanini represented a pair of 5S rDNA signals equally. In addition, F. tataricum exhibited two pairs of 45 rDNA signals, and F. tataricum ssp. potanini showed a pair of 45S rDNA signals. Our results were consistent with previous studies suggesting that F. tataricum evolved from F. tataricum ssp. potanini. Additionally, we found that F. tataricum ssp. potanini has the fewest rDNA repeats in the cymosum group. These results can be used as basic cytological information for buckwheat research in the future.

Karyotype Report
  • Lizhu Qian, Sven Landrein, Chunhui Hao, Fuchuan Wu
    2023 Volume 88 Issue 4 Pages 295-300
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Six Passiflora species from SW China were studied: Passiflora menghaiensis X.D.Ma, L.C.Yan & J.Y.Shen, P. xishuangbannaensis Krosnick, P. siamica Craib, P. altebilobata Hemsl., P. henryi Hemsl. and P. papilio Li. Chromosomes squashes could be obtained by using the conventional tableting method and the karyotypes were analyzed. The results showed that all six species of passion flower were diploid and the chromosome number was 2n=12. The karyotype formula of P. menghaiensis was 2n=2x=12=8m+2sm+2sm(sat), P. xishuangbannaensis was 2n=2x=12=8m+4sm, P. siamica was 2n=2x=12=10m+2sm, P. altebilobata was 2n=2x=12=10m+2sm, P. henryi was 2n=2x=12=8m+4sm, P. papilio was 2n=2x=12=12m. The karyotypes of all six species are reported for the first time. This study provides comprehensive cytological data for the conservation and genetic study of Passiflora in Asia. The evolutionary relationships between the six species were discussed at the cytological level and compared with published classifications based on morphological and molecular data.

  • Komal Arora, Minakshi Sharma, Rajneesh Kant Sachdev, Lovleen
    2023 Volume 88 Issue 4 Pages 301-306
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Eucalyptus citriodora Hook., commonly known as lemon-scented eucalypt, is an aromatic plant of the family Myrtaceae. The present study was planned to evaluate the genotoxic and cytotoxic activity of Eucalyptus citriodora essential oil (ECEO) using Allium cepa as a model plant. This essential oil extracted from leaves of E. citriodora was analyzed using a Headspace GC-MS system. Citronella, β-Citronellol, Isopulegol, and Citronellyl acetate were the major components in ECEO. Mitotic studies in A. cepa root tip cells showed a dose-dependent increase in chromosomal aberration and a decrease in mitotic index after 24 h treatment with various concentrations of ECEO. Main chromosomal abnormalities included sticky metaphase, metaphase fragmentation, vagrant chromosome, and C-mitosis. Chromosomal stickiness was the most observed chromosomal alteration in all treatments. Thus, it confirmed the cytotoxic and genotoxic effects of ECEO on A. cepa. The study concludes that ECEO has inhibitory potential toward the growth of other plants which can be exploited to develop eco-friendly bio-herbicides.

  • Rohita Singla, Harcharan Singh Dhaliwal, Vivek Sharma
    2023 Volume 88 Issue 4 Pages 307-311
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    One of a common commercial and culinary fruits from the Solanaceae family is the Cape gooseberry. Due to their enormous medicinal potential to treat a variety of ailments, the Physalis species have been employed traditionally. The Physalis species has a wide range of bioactivities based on the presence of secondary metabolites, including immunomodulatory, cytotoxic, anti-diabetic, antioxidant, anti-cholesterolemic, and antimicrobial. The plants gathered from various parts of Himachal Pradesh are subjected to meiotic exams in this study. The purpose of the current research was to study the ploidy levels of the aforementioned Physalis species using conventional chromosome counting techniques. More cytotypes, morphotypes, and chemotypes must be identified in order to conduct future breeding experiments for getting a crop with high medicinal content and stress tolerant variety. Therefore, morphological and meiotic analyses of three Indian Physalis species were performed. P. angulata L. (2n=4x=48) demonstrates a tetraploid nature, whereas P. peruviana L. (2n=2x=24) and P. longifolia Nutt. (2n=2x=24) are diploid.

  • Jia Miao, Xin-Fen Gao, Zhen-Long Liang, Xue-Li Zhao
    2023 Volume 88 Issue 4 Pages 331-337
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Supplementary material

    The genera Uraria Desv. and Christia Moench are sister genera belonging to the tribe Desmodieae (Fabaceae), and most species are used as important medicinal or ornamental plants. Evolutionary patterns and intergeneric delimitation of these genera have long been controversial. In the present study, the chromosome numbers and karyotypes of six Uraria species and three Christia species were analyzed via the root tip chromosome squashing method towards a better understanding of karyotype evolution. The chromosome numbers observed in Uraria and Christia were 2n=2x=22 and 2n=2x=20, respectively. Analysis of karyotypes resulted in formulas of 20m+2sm (U. crinita and U. lacei), 18m+4sm (U. lagopodioides), 22m (U. picta, U. rufescens, and U. sinensis), and 20m (C. campanulata, C. obcordata, and C. vespertilionis). Cluster analysis conducted using the UPMGA algorithm based on chromosomal and karyotypic parameters revealed that these nine species were divided into two clades, where species of Uraria and Christia each formed a monophyletic clade. In the Uraria clade, U. picta, U. sinensis, U. crinita, and U. rufescens were divided into a subclade, while U. lacei and U. lagopodioides were divided together, showing their close genetic relationships. The taxonomically controversial species, C. campanulata, nested within the Christia clade and showed a close affinity to C. vespertilionis.

  • Murat Kürşat, Osman Gedik, Irfan Emre
    2023 Volume 88 Issue 4 Pages 347-352
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    The goal of this study is to conduct a thorough karyotypic examination of nine Ranunculus species [R. brachylobus subsp. incisilobatus, R. kotschyi (B1), R. kotschyi (B2), R. macrorrhynchus subsp. trigonocarpus, R. polyanthemos, R. repens, R. sericeus, R. sphaerospermus, and R. strigillosus] growing in Turkey. This study discovered that the basic number of Ranunculus taxa investigated was x=7 or 8. R. sphaerospermus and R. strigillosus have median and submedian chromosomes while other taxa have subterminal (st) chromosomes as well median (m) and submedian (sm). This investigation also revealed that R. kotschyi (B1), R. kotschyi (B2), and R. macrorrhynchus subsp. trigonocarpus are hexaploid, whereas R. repens and R. sphaerospermus are tetraploid. The Stebbins classification, ploidy levels, karyotype formula, chromosomal length range, total karyotype length, various karyotype asymmetry values, and the A1 and A2 in each taxon were also determined in this study. The Stebbins classification of R. brachylobus subsp. incisilobatus, R. polyanthemos, and R. repens were 3A while the other six taxa have 3B.

  • Rupali N. Chougule, Nilesh V. Pawar, Mansingraj S. Nimbalkar, Manoj M. ...
    2023 Volume 88 Issue 4 Pages 373-377
    Published: December 25, 2023
    Released on J-STAGE: December 25, 2023
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    Lavandula gibsonii is an endemic lavender that occurs in the Western Ghats of Maharashtra and Tamil Nadu states of India. It is aromatic and has insecticidal properties. We studied the cytogenetics of the species for the first time. The chromosome number was observed to be 2n=40 and the small chromosomes ranged from 0.95 to 2.05 µm. Meiosis was normal with n=20 bivalents at diakinesis. Somatic metaphase chromosomes with submedian region centromeres dominated the karyotype. The karyotype formula was 2m + 18sm. The karyotype was found to be asymmetrical and fell into Stebbins’ 4B category. The base number (x) of section Chaetostachys in which Indian species are placed needs to be reassessed.

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