• Journal of the Korean Society of Tobacco Science
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• v.20 no.1
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• pp.57-65
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• 1998

This study was conducted to develop a resistant tobarro against Potato virus Y (PVY) by transformation of the plants with genetically engineered viral genes. The complimentary DNAs (cDNAS) of potato virus Y-necrosis strain (PVY-Vn) replicase mutant genes (3'-deleted, 5'-deleted and ADD-mutant Nlbs) were synthesized through RT-PCR by using purified PVY-VN RNA and synthesized primers, and cloned in the sense orientation into a plant expression vector (pMBPI), The cDNAS of the genes were transferred into Agrobacterium tumefaciens LBA 4404, and then transformed into tobacco (Nicotiana tabacum cv. Burley 21) plants. Regenerated plants were tested for PVY resistance by inoculation test; 13 transgenic plants including 7 for 3'-deleted Nlb, 3 for 5'-deleted Nlb, and 3 for ADD-mutant Nlb appeared to be resistant at 4 weeks after inoculation with PVY-VN. Among the 13 transgenic tobacco plants, 8 plants had no symptom up to 14 weeks after inoculation. The progenies ($T_1$) from self-fertilization of the transgenic lines varied 0.0% to 81.2% in their resistance (% of resistant plants). The analysis of Nlb-31deleted, -5'deleted and -ADD mutant in the $T_1$ plants by polymerase chain reaction (PCR) showed that Nlb-3'deleted, -5'deleted and -ADD mutants were detected in all of the resistant plants. These results suggest that the PVY resistance was inherited in the $T_1$ generation.

• Korean Journal of Plant Resources
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• v.11 no.2
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• pp.188-194
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• 1998

This experiment was conducted to introduce phosphinothricin acetyl -transferase(PAT) gene, resistant to basta and non-selective herbidide, into tobacco(Nicotiana tabacum cv.BY4). For shoot formation,tobacco leaf disks were placed on the MS medium supplemented with 2.0mg/L BA and 0.1mg/L NAA. In this medium condition, tobacco leaf disces were cocultivated with A. tumefaciens MP90 containing NPT IIand PAT resistant to kanamycin and Basta, respectively. Shoots were obtained in the medium containing antibiotics, and those were transferred to rooting medium supplemented with 0.1mg/L NAA and antibiotics. The plants obtaining roots were transplanted into soil. Phenotype of transgenic tobacco plant was mostly as normal plant. However, about 5% was abnormal plant, which did not set seeds. PCR analysis and southern blot were performed to determine transformation. As the results, it was confirmed that PAT gene was stably integrated into tobacco genome.When herbicide, basta, was sprayed to the plants confirmed by PCR, the transgenic plants showed normal growth, whereas normal plants died. Therefore, the result of this experiment show that tobacco transformation for the resistance to basta, non-selective herbicide, was successful because PAT gene was stably integrated into tobacco.

• Journal of the Korean Society of Tobacco Science
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• v.6 no.2
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• pp.185-189
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• 1984

To classify the inheritance of resistance to potato virus Y, crosses between susceptible flue-cured tobacco variety NC 95 and resistant variety McNair 30 were conducted. The parents, $F_1$ plants, $F_2$ populations, and haploid plants derived from anthers of $F_1$ plants were screened for a resistance of two potato virus Y strains (PVY-VB and PVY-VN) isolated in Korea. The Chi-square values for the $F_3$ populations and haploids of $F_1$ fitted 1 :3 and 1 :1 ratios of resistant to susceptible for two strains, respectively. Therefore, it was found that the resistance of McNair 30 for the potato virus Y was controlled by a single recessive gene. Moreover the resistance to two strains screened was inherited dependently.

• Journal of the Korean Society of Tobacco Science
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• v.19 no.2
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• pp.83-91
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• 1997

The vein-necrosis strain or potato virus Y (PVY-Vff) and black shank (Phytophlhora parasitica roar. nicotianae) causes severe damage on burley tobacco(Wicotiana tabacum L.) in Korea, A new burley tobacco resistance to PVY and black shank, KB 110, was developed by Korea Ginseng and Tobacco Research Institute. It was developed from the cross of Burley 21 with TC 591 in 1990, and was backrossed to Burley 21 in the following season. TC 591 has resistance to PVY and moderate resistance to race 0 of black shank, but it is susceptible to tobacco mosaic vim (TMV). KB 110 was evaluated for its resistance to PVY, TMV and black shank in the greenhouse and at fields for preliminary and performance trials. KB 110 which has secreting glandular trichomes was resistant to PVY-VN, TW and black shank. It had an erect growth habit and two more leaves per plant than that of Burley 21, and matures two to three days later. It yielded approximately 3 percent more cured leaf than the standard cultivar Burley 21, but other plant characteristics were very similar to those of Burley 21. It had acceptable standards for chemical and physical characteristics of lured leaf on regional farm test in 1995-1997. KB 110 produced average yields of good quality tobaccos and was appeared to be resistant to PVY inwhere occurrence of the virus are severe chronic at burley growing area.

• BMB Reports
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• v.31 no.3
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• pp.287-295
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• 1998

Acetolactate synthase (ALS) is the first common enzyme in the biosynthesis of L-Ieucine, L-isoleucine, and L-valine. The sulfonylurea-resistant ALS gene from Nicotiana tabacum was cloned into the bacterial expression vector pGEX-2T. The resulting recombinant plasmid pGEX-ALS3 was used to transform Escherichia coli strain XL1-Blue, and the mutant tobacco ALS (mALS) was expressed in the bacteria as a protein fused with glutathione S-transferase (GST). The fusion product GST-mALS was purified in a single step on a glutathione-Sepharose column. ALS activities of 0.9-2.5 ${\mu}mol/min/mg$ protein were observed in the GST-mALS, and the Km values for pyruvate, FAD, and TPP were 10.8-24.1, $(1.9-8.9){\times}10^{-3}$, and 0.14-0.38 mM, respectively. The purified GST-mALS was resistant to both the sulfonylurea and the triazolopyrimidine herbicides, and lost its sensitivity to end products, L-valine and L-leucine. For comparision, the tobacco wild-type recombinant ALS fused with GST, GST-wALS, was also characterized with respect to its pyruvate and cofactor bindings. These results suggest that the purified mutant recombinant tobacco ALS was functionally active, that the mutations resulting in herbicide resistance has affected pyruvate and cofactor bindings," and that the two classes of herbicides interact at a common site on the plant ALS.

• The Plant Pathology Journal
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• v.16 no.5
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• pp.264-268
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• 2000

A fluorescent material was accumulated in inoculated leaves showing necrotic local lesions of tobacco plants with N gene, Nicotiana tabacum cvs. Xanthi-nc NN, Samsun NN, Burley 21 and KF 114, and N. glutinosa, and Datura stramonium at the early growth stages by the inoculation of Tobacco mosaic virus (TMV). It was identified as a coumarin phytoalexin, scopoletin. Although the material was most prominently produced in TMV-inoculated tobacco leaves with local necrotic lesions, its accumulation was also noted in uninoculated leaves of TMV-inoculated plants. Its accumulation was somewhat greater in high resistance-induced leaves than low resistance-induced and intact leaves. Scopoletin treatment induced the expression of a pathogenesis-related protein, PR-1, prominently at the concentration of 500 or 1000 ${\mu}$g/ml. This suggests that scopoletin is a phytoalexin abundantly accumulating in N gene-containing resistant plants in response to TMV infection, and may be related to hypersensitive responses (HR) and systemic acquired resistance (SAR) in the resistant tobacco plants.

• Proceedings of the Korean Society of Tobacco Science Conference
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• 1997.10a
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• pp.134-152
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• 1997

Plant viruses of tobacco including tobacco mosaic virus (TMV) and potato virus Y (PVY) cause severe economic losses in leaf-tobacco production. Cultural practices do not provide sufficient control against the viruses. Use of valuable resistant cultivars is most recommendable for the control of the viruses. However, conventional breeding programs are not always proper for the development of virus-resistant plants mostly owing to the frequent lack of genetic sources and introduction of their unwanted properties. Therefore, we tried to develop virus-resistant tobacco plants by transforming commercial tobacco cultivars, NC 82 and Burley 21, with coat protein (CP) or replicase (Nlb) genes of TMV and PVY necrosis strain (PVY-VN) with or without untranslated region (UTR) and with or without mutation. Each cDNA was cloned and inserted in plant expression vectors with 1 or 2 CaMV 35S promotors, and introduced into tobacco leaf tissues by Agrobacterium tumefaciens LBA 4404. Plants were regenerated in kanamycin-containing MS media. Regenerated plants were tested for resistance to TMV and PVY In these studies, we could obtain a TMV-resistant transgenic line transformed with TMV CP and 6 genetic lines with PVY-VN cDNAs out of 8 CP and replicase genes. In this presentation, resistance rates, verification of gene introduction in resistant plants, stability of resistance through generations, characteristics of viral multiplication and translocation in resistant plants, and resistance responses relative to inoculum potential and to various PVY strains will be shown. Yield and quality of leaf tobacco of a promising resistant tobacco line will be presented.

• Research in Plant Disease
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• v.8 no.2
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• pp.78-83
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• 2002

Tobacco diseases have not been recorded until 1900s in Korea, where tobacco plants were introduced at early 1700s. Practical researches on the disease have been conducted since mid 1960s. Major ten tobacco diseases were mosaic caused by tobacco mosaic virus·potato virus Y·cucumber mosaic virus, bacterial wilt, hollow stalk, wild fire caused by angular leaf spot strain, black shank, brown spot, powdery mildew and fusarium wilt. But their annual occurrences were varied according to changes of tobacco varieties and their cultivating practices. As no useful chemicals, several biological tactics have been developed to control the viral or bacterial diseases that give significant economic damages on sustainable crop yield, but not practicable to field farming condition yet. Transgenic tobacco plants containing foreign disease resistant genes have been developed by current bio-technology, but not released to farmers yet. Though some disease-resistant tobacco varieties have been developed by the conventional breeding technology and currently used by farmers, their disease controlling efficacy have been diminished by occurrence of the new strain or race. Future research on tobacco diseases has been focused on technical development to produce high quality tobacco with less production cost, which leads Korean tobacco industry to keep its competence against foreign industry and decreasing overall market.

• Korean journal of applied entomology
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• v.23 no.2 s.59
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• pp.96-101
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• 1984

This experiment was carried out to investigate control efficacy by cultural practice and fumigation of tacterial wilt caused by Pseudomonas solancearum in resistant tobacco line NC82 at the Korea Ginseng and Tobacco Research Institute, Eumseong Experiment Station in 1982 and 1983. The bacterial wilt of tobacco occurred severely from mid-July to last August in applicable temperature and soil humidity for increasing bacteria. Disease severity appeared low and slowly at fumigation and resistant variety treatment. Incidence of bacterial wilt in tobacco line NC82 was $44.7\~55.8\%$ being compared with susceptible variety, NC2326 and $95\~99\%$ when resistant variety, NC32 was cultivated with soil fumigation treatment. Control efficacy of cultural practices appeared low with $0.8\~20\%$ and was not different from resistant variety and fumigation treatment. Control system against bacterial wilt in flue-cured tobacco was accomplished by control efficacy over $95\%$ when resistant variety(NC82) was cultured after treatment of cultural practices (Tillering after harvest and before transplanting, stalk and root destruction, early transplanting early removal of the mulching film) and soil fumigation(Cylon).

• Research in Plant Disease
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• v.9 no.4
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• pp.224-228
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• 2003

Five commercial chemicals, Dimethomorph, Foseyl-Al, Oxadixyl + Mancozeb, Propamocarb hydrochloride, and Metalaxyl, were tested for control of tobacco black shank (Phytophthora nicotianae) and/or delaying buildup of the resistant population to Metalaxyl which has been used for control of the disease in Korean tobacco farms. Propamocarb hydrochloride seemed to have a cross-resistance to the Metalaxyl resistant isolates showing similar response with Metalaxyl in vitro. Meanwhile, Dimethomorph+Mancozeb and Oxadixyl+Mancozeb were selected to be the promising chemicals which are able to be alternative to Metalaxyl for the black shank control in accordance with in vitro and in field trial.