• Korean Journal of Plant Tissue Culture
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• v.22 no.4
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• pp.195-200
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• 1995

The mammalian adenosine deaminase(ADA) gene was stably expressed in transgenic tobacco plane. The chimeric ADA gene 35S/35S/AMV/ADA/Tnos, has been constructed. This chimeric gene was introduced into the binary vector pRD400, which was thereafter mobilized into Agrobacterium tumefaiens strain MP90 harboring disarmed Ti-plasmid. The resulting strains were used to transform Nicofiana tabacum L. using the leaf disc. Incorporation of the chimeric gene into plant were confirmed by PCR and Northern blot analyses. Immunoblot analysis showed that ADA protein was successfully synthesized in the transgenic tobacco plants.

• Molecules and Cells
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• v.44 no.12
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• pp.911-919
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• 2021

The virus-induced genome editing (VIGE) system aims to induce targeted mutations in seeds without requiring any tissue culture. Here, we show that tobacco rattle virus (TRV) harboring guide RNA (gRNA) edits germ cells in a wild tobacco, Nicotiana attenuata, that expresses Streptococcus pyogenes Cas9 (SpCas9). We first generated N. attenuata transgenic plants expressing SpCas9 under the control of 35S promoter and infected rosette leaves with TRV carrying gRNA. Gene-edited seeds were not found in the progeny of the infected N. attenuata. Next, the N. attenuata ribosomal protein S5 A (RPS5A) promoter fused to SpCas9 was employed to induce the heritable gene editing with TRV. The RPS5A promoter-driven SpCas9 successfully produced monoallelic mutations at three target genes in N. attenuata seeds with TRV-delivered guide RNA. These monoallelic mutations were found in 2%-6% seeds among M₁ progenies. This editing method provides an alternative way to increase the heritable editing efficacy of VIGE.

• Korean Journal Plant Pathology
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• v.13 no.3
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• pp.179-183
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• 1997

We have screened hypersensitive responses of 18 cultivars of Nicotiana tabacum to Xanthomonas campestris pv. campestris. TC500 cultivar produced the most strong hypersensitive response (HR) to Xanthomonas campestris pv. campestris. By NTG mutagenesis, nonhypersensitive mutants (XHN 514-774, XHN 620-831) were generated, which does not induce hypersensitive response on tobacco leaves (Nicotiana tabacum cv. TC500). Also nonpathogenic mutant (XPN 1001), which does not incite any of the black rot symptoms on leaves was generated. We observed that HR mutants were still pathogenic on cabbage leaves producing black rot symptoms and nonpathogenic mutant induced HR in tobacco leaves. The inplanta growth of wild type and HR mutants were examined for up to 120 hrs after inoculation : population of wild type strain increased to $10^{8}$ in 24hrs, but rapidly declined thereafter; HR-mutants increased to more than $10^{6}$ in 48 hrs after inoculation but subsequently stabilized and slowly decreased. We observed that wild type and these mutans produced similar amounts of degradative enzymes such as protease, pectate lyase, cellulase and amylase.

• Korean Journal of Plant Resources
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• v.18 no.3
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• pp.359-366
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• 2005

Effects of neutron beam irradiation on seed germination, growth and RAPD pattern of tobacco (Nicotiana tabacum L. cv.; N. plumbaginifolia) and rice (Orya sativa L. cv.) plants were estimated. Seed germination rate was not significantly changed by the neutron beam treatment in both tobacco and rice seeds. And there was no significant differance in growth of the plants by the neutron beam treatment. Interestingly, however, some of morphological changes, including leaf shape (about $36\%$), stem color and leaf color were observed in neutron beam treated tobacco plants. In addition, abnormal flower in petal was observed in the neutron beam treated plant. This results indicate that neutron beam is able to use as an effective mutagen in plant mutations. Scorable products from 20 primers were obtained by RAPD analysis in the leaves of the beam irradiated tobacco plants and most of the plants showed the similar band patterns.

• Research in Plant Disease
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• v.12 no.3
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• pp.226-230
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• 2006

Cucumber mosaic virus(CMV) was occurred on red pepper showing vein chlorosis or vein necrosis with the incidence rate of 52% from 62 specimens collected in natural fields. Among 32 samples infected with CMV, the specimens of 22 red pepper leaves showing vein chlorosis were infected singly with CMV-VCH. CMV-VCH induced vein chlorosis on the inoculated leaves, and vein banding and vein necrosis on the upper leaves of Nicotiana glutinosa, and then killed after showing bud necrosis. The typical symptoms of vein banding, malformation and blister were produced on the upper leaves of Nicotiana benthamiana and N. tabacum 'Ky-57' without symptoms on the inoculated leaves. The commercial cultivars of 'Bugang', 'Manitta' and 'Gwariput' were shown the typical symptom of vein chlorosis by the mechanical inoculation of CMV-VCH. CMV-VCH was detected specifically by RT-PCR. Virus particles of CMV-VCH were isometric shape having 30 nm diameter. Ultraviolet absorption of purified CMV-VCH was maximum at 260 nm and minimum at 242 nm. The ratio of A260/A280 was 1.71. CMV-VCH had the single nucleo-protein having the molecular weight of 24.5 kDa.

• Korean Journal of Plant Tissue Culture
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• v.27 no.2
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• pp.109-115
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• 2000

Effects of the heavy-ion ($^{14}$ N or $^{20}$ Ne) beam irradiation on the response of anthers, growth of calli, germination of seeds, and the early growth after the germination of tobacco (Nicotiana tabacum L. cv. BY-4) were investigated. Anthers precultured for 10 days before the irradiation became brown without callus or shoot induction over 20 Gy of $^{14}$ N and $^{20}$ Ne ion beam irradiation. Relative growth rate of the cultured calli was reduced by the irradiation and became brown significantly 2 weeks after the $^{14}$ N and $^{20}$ Ne ion beam irradiation over 50 Gy. The increased intensity of the heavy-ion ($^{14}$ N, $^{20}$ Ne) beam irradiation resulted in the delay of seed germination and the inhibition of the early growth both in water-treated and non-treated seeds before the irradiation. In addition, the heavy-ion beam irradiation to the imbibed seeds inhibited seed germination more than that to the non-imbibed seeds. The screening approach of non-imbibed seeds with heavy-ion beam irradiation using in vitro culture system was more useful than the filter-paper germination method in investigating the characteristics of heavy-ion beam-irradiated seed population and the screening of morphological variants at the early stage of the plant growth.

• Korean Journal of Plant Tissue Culture
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• v.21 no.1
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• pp.23-28
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• 1994

This study was rimed out to examine the difference in the cell vitality between mesophyII protoplast (MP) and paraveinal mesophyII protoplast (PVMP) of Nicotiana tabaccum 'Xanti', Petunia hybrida 'Blue Star' and Chrysanthemum morifolium 'Baeckwang' by using urea permeability technique. The effects of various enzyme solutions and incubation time, NAA and thidiazron on plant regeneration from isolated protoplasts were also investigated. The vibratome technique was used for protoplast isolation and urea permeability test because the fresh living, thin tissue stripes (50 ${\mu}{\textrm}{m}$ of thickness) could be obtained with minimal damage with the vibratome. For the three plants examined, the urea permeability on the tested tissue stripes was relatively higher in PVMP than in MP by about Ks = 2.0 $\times$ 10$^{-5}$ cm/sec. The treatment of an enzyme mixture of 1.5% cellulase R-10, 1% Driselase, 0.5% Macerozyme R-10, and 0.5% Pectinase for 4 to 8 h was effective on the isolation of PVMP. The highest frequency of callus formation and plant regeneration from the isolated protoplasts was obtained with NAA 2 mg/L and thidiazuron 0.01 mg/L. Furthermore, the results demonstrated that cell devision and plantlet regeneration was more frequent in the PVMP than in the MP of the same leaf or plant We, therefore, conclude that UM is an excellent experimental material for the callus formation and regeneration from isolated protoplasts.

• Applied Biological Chemistry
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• v.37 no.5
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• pp.343-348
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• 1994

Ti plasmid of A. tumefaciens was labeled with $^3H-thymidine$, purified and encapsulated into phosphatidylserine (PS) and PS-cholesterol (Chol; 1 : 1 molar ratio) liposomes by lyophilization-rehydration method. PS was supplemented with 1 mole percent octadecyl rhodamine B for fluorometric measurement of PS. Liposomes entrapping $^3H-Ti plasmid$ were fused with Nicotiana sanderae protoplasts by treating with 5 mM $CaCl_2$ and 10% PEG. The fusion was evidenced by fluorescence microscopic technique. The amounts of Ti plasmid and PS associated with protoplasts were assayed by the radioactivity of $^3H-Ti plasmid$ and by the fluorescence of rhodamine B. About 7.9% of the PS liposome and 7.2% of PS-Chol liposome were fused with protoplasts. During the fusion process, about 30% of the liposomal contents of PS-Chol liposome was leaked, in contrast to about 60% leakage of its contents in PS liposome. Accounting the number of liposomes fused with protoplasts together with the encapsulation efficiency and the leakage of liposomal contents, it was calculated that ca. 1,700 Ti plasmid was transfered into one protoplast by the present method. This result may indicates that the present method transfers enough Ti plasmid into plant protoplast to elicit genetic transformation of plants.

• Journal of Plant Biotechnology
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• v.47 no.4
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• pp.337-344
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• 2020

Plant growth-promoting microorganisms promote plant growth by supplying nutrients to roots and interacting with the intrinsic factors in plants through volatile organic compounds (VOCs). In this study, we evaluated the effect of UOS, plant growth-promoting fungi (PGPF) isolated from previous study, on the growth of Nicotiana tabacum L. var Xanthi nc. Phylogenetic analysis and GC-MS were used to identify the fungal species and the VOCs emitted by the UOS, respectively. The fresh weight of UOS-treated Nicotiana tabacum L. was 3.8 and 4.2-fold higher than that of the control groups grown in vertical and I-plates, respectively. Moreover, in the UOS-treated plants, the length of the primary root was half and the number of lateral roots were twice compared to those in control plants. The UOS was identified as Phoma sp. by studying spore and mycelial morphology and using phylogenetic analysis. GC-MS revealed that the VOC emitted by the UOS was hexamethylcyclotrisiloxane (D3). These results suggest that the UOS of Phoma sp. influences plant growth and root development through D3. We expect this UOS and its VOC, D3 to be utilized in the future to increase growth and enhance yield for other plants.

• Journal of Ecology and Environment
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• v.45 no.3
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• pp.143-151
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• 2021

Background: Plants are able to optimize defense responses induced by various herbivores, which have different feeding strategies. Local and systemic responses within a plant after herbivory are essential to modulate herbivore-specific plant responses. For instance, leaf-chewing herbivores elicit jasmonic acid signaling, which result in the inductions of toxic chemicals in the attacked leaf (tissue-specific responses) and also in the other unattacked parts of the plant (systemic responses). Root herbivory induces toxic metabolites in the attacked root and alters the levels of transcripts and metabolites in the unattacked shoot. However, we have little knowledge of the local and systemic responses against stem-boring herbivores. In this study, we examined the systemic changes in metabolites in the wild tobacco Nicotiana attenuata, when the stem-boring herbivore Trichobaris mucorea attacks. Results: To investigate the systemic responses of T. mucorea attacks, we measured the levels of jasmonic acid (JA), JA-dependent secondary metabolites, soluble sugars, and free amino acids in 7 distinct tissues of N. attenuata: leaf lamina with epidermis (LLE), leaf midrib (LM), stem epidermis (SE), stem pith (SP), stem vascular bundle (SV), root cortex with epidermis (RCE), and root vascular bundle (RV). The levels of JA were increased in all root tissues and in LM by T. mucorea attacks. The levels of chlorogenic acids (CGAs) and nicotine were increased in all stem tissues by T. mucorea. However, CGA was systematically induced in LM, and nicotine was systematically induced in LM and RCE. We further tested the resource allocation by measuring soluble sugars and free amino acids in plant tissues. T. mucorea attacks increased the level of free amino acids in all tissues except in LLE. The levels of soluble sugars were significantly decreased in SE and SP, but increased in RV. Conclusions: The results reveal that plants have local- and systemic-specific responses in response to attack from a stem-boring herbivore. Interestingly, the level of induced secondary metabolites was not consistent with the systemic inductions of JA. Spatiotemporal resolution of plant defense responses against stem herbivory will be required to understand how a plant copes with attack from herbivores from different feeding guilds.