• Journal of Applied Biological Chemistry
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• v.43 no.2
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• pp.69-73
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• 2000

In order to understand the biological role of calmodulin in plants, transgenic plants expressing a mutant calmodulin (VU-4, Iys to ile-115) have been analyzed. We found that tobacco plants expressing VU-4 calmodulin have approximately twofold higher $\gamma$-aminobutyric acid (GABA) levels than the control plants. Cell suspension cultures established from the stem explants of the transgenic tobacco seedlings also have higher levels of GABA than the control cell cultures. Specific activity of glutamate decarboxylase (GAD), which catalyzes the decarboxylation of glutamate to $CO_2$ and GABA, of the transgenic tobacco cell extracts was about twofold higher than the activity of the control cell extracts. Western-blot analysis showed that the GAD is highly expressed in the transgenic tobacco plants. GAD partially purified from tobacco cell extracts showed approximately threefold $Ca^{2+}$/calmodulin-dependent activation. These data suggest that GABA synthesis in the transgenic tobacco plants is elevated, possibly due to higher levels of the calmodulin-dependent GAD enzyme and/or as a result of enhanced activation due to increased levels of the foreign calmodulin.

• Biotechnology and Bioprocess Engineering:BBE
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• v.11 no.2
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• pp.154-159
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• 2006

Interleukin-18 (IL-18), otherwise known as interferon-gamma-inducing factor (IGIF), is one of several well characterized and important cytokines that contribute to host defenses. The complementary DNA (cDNA) of mature human interleukin-18 gene (hIL-18) was fused with the signal peptide of the rice amylase 1A gene (Ramy1A) and introduced into the plant expression vector under the control of a duplicated CaMV 35S promoter. The recombinant plasmid was transformed into tobacco (Nicotiana tabacum L. cv Havana) using the Agrobacterium-mediated transformation method. The integration of the hlL-18 gene into the genome of transgenic tobacco plants was confirmed by polymerase chain reaction (PCR) amplification and its expression was observed in the suspension cells that were derived from the transgenic plant callus by using Northern blot analysis. The hlL-18 protein was detected in the extracts of the transgenic callus and in the medium of the transgenic tobacco suspension culture by using immunoblot analysis. Based upon enzyme-linked immunosorbant assay (ELISA) results, the expression level of the hlL-18 protein approximated $166{\mu}g/L$ in the suspension culture medium. Bioassay results from the induction of $interferon-{\gamma}$ from a KG-1 cell line indicated that the hlL-18 secreted into the suspension culture medium was bioactive.

• Molecules and Cells
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• v.21 no.1
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• pp.141-146
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• 2006

We previously showed that NtCDPK1, a tobacco calcium-dependent protein kinase, interacts with and phosphorylates the Rpn3 regulatory subunit of the 26S proteasome, and that both NtCDPK1 and Rpn3 are mainly expressed in rapidly proliferating tissues, including shoot and root meristem. In this study, we examined NtCDPK1 expression in roots using GUS expression in transgenic Arabidopsis plants, and investigated its function in root development by generating transgenic tobacco plants carrying a sense NtCDPK1 transgene. GUS activity was first detected in roots two days after sowing. In later stages, strong GUS expression was detected in the root meristem and elongation zone, as well as the initiation sites and branch points of lateral roots. Transgenic tobacco plants in which NtCDPK1 expression was suppressed were smaller, and their root development was abnormal, with reduced lateral root formation and less elongation. These results suggest that NtCDPK1 plays a role in a signaling pathway regulating root development in tobacco.

• Journal of Plant Biotechnology
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• v.7 no.2
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• pp.97-103
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• 2005

This study was carried out to obtain basic information for gene manipulation in potent edible tobacco (Nicotiana tabacum cv. TI 516). N. tabacum cv. TI 516 is a plant for a possible candidate to use as an edible vaccine, since it contains a low level of nicotine. The effective plant regeneration system through leaf disc culture was achieved using a MS basal medium supplemented with 0.1 mg $1^{-1}$ NAA and 0.5 mg $1^{-1}$ BA. In order to transform the N. tabacum cv. TI 516 with the green fluorescent protein (GFP) gene, Agrobacterium tumefaciens LBA 4404 containing the GFP gene was used. Genomic PCR confirmed the integration of the GFP gene into nuclear genome of transgenic plants. Expression of the GFP gene was identified in callus, apical meristem and root tissue of transgenic N. tabacum cv. TI 516 plants using fluorescence microscopy. Western blot analysis revealed the expression of GFP protein in the transgenic edible tobacco plants. The amount of GFP protein detected in the transgenic tobacco plants was approximately 0.16% of the total soluble plant protein (TSP), which was determined by ELISA.

• 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 Environmental Science International
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• v.10 no.1
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• pp.79-84
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• 2001

To assess resistance of transgenic tobacco plants which overexpress superoxide dismutase (SOD) and ascorbate peroxidase (APX) in chloroplasts to water stress, changes in leaf water potential, turgor potential, stomatal conductance and transpiration rate were measured. Leaf water potential in all plants remained high up to day 4 after withholding water but thereafter decreased markedly. In spite of a remarkable decrease in leaf water potential, some of transgenic plants maintained higher turgor potential compared with control plant on day 12. In particular, the transgenic plant expressing MnSOD showed an outstanding maintenance in turgor pressure by osmotic adjustment throughout the experiment, resulting in high stomatal conductance and transpiration rate. However, among transgenic plants, osmotic potential was reduced more effectively in multiple transformants such as the double transformant expressing both MnSOD and APX, and the triple transformant expressing CuznSOD, MnSOD and APX than single transformants. Consequently, further research is needed to get general agreement on the tolerance of transgenic plants to water stress at different growth stages for each transgenic plant.

• Journal of the Korean Society of Tobacco Science
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• v.25 no.1
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• pp.12-19
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• 2003

Transgenic tobacco plants were selected by using the transformation of putrescine N-methyltransferase(PMT) gene, the key enzyme in diverting polyamine metabolism towards the biosynthesis of nicotine. PMT was fused in reverse orientation to the CaMV 35S promoter of the plant expression vector pBTEX(pPAB3) to produce tobacco plants of low nicotine content. To compare nicotine content, only pBTEX vector and PMT gene which was fused in forward orientation to the CaMV 35S promoter(pPAB2) were also transformed to the leaf tobacco plants(Nicotiana tabacum cv. NC82 and N. tabacum cv. Br2l). The presence of sense- and antisense-PMT gene, and pBTEX vector in the transgenic plant was confirmed by genomic PCR.

• Journal of Plant Biotechnology
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• v.43 no.4
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• pp.432-437
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• 2016

Interleukin-11 (IL-11) is a cytokine that plays a key regulatory role in the immune system. Recombinant human IL-11 (rhIL-11) exerts a preventative effect against apoptotic cell death and inhibits preadipocyte differentiation. IL-11 also is used to stimulate the bone marrow to produce platelets in order to prevent low platelets that may be caused by chemotherapy. Unfortunately, the high production cost of IL-11 associated. In this study, we investigated the feasibility of transgenic plants for the cost-effective production of rhIL-11. Production of rhIL-11 proteins in whole-plant expression system will be more economical when compared to the current E. coli based expression system. The human rhIL-11 gene was codon optimized to maximize plant host system expression. IL-11 expression vector under the control of a constitutive cauliflower mosaic virus 35S (CaMV 35S) promoter was introduced into tobacco by Agrobacterium-mediated transformation. The 5'-leader sequence (called ${\Omega}$) of tobacco mosaic virus (TMV) as a translational enhancer was added to construct. Transgenic tobacco plants expressing various levels of rhIL-11 protein were generated. Western blotting of the stably transformed lines demonstrated accumulation of the appropriately sized rhIL-11 protein in leaves. This research demonstrated the efficacy of using tobacco as an expression system for the production of rhIL-11.

• Proceedings of the Botanical Society of Korea Conference
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• 2001.04a
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• pp.23-23
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• 2001

• Journal of Plant Biotechnology
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• v.35 no.4
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• pp.329-335
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• 2008

Tocopherols are essential lipophilic antioxidant in human cells, while little is known about its function in plant tissues. To study the impact of composition and content of tocopherols on stress tolerance, tobacco (Nicotiana tabacum) was transformed with a construct containing a cDNA insert encoding $\gamma$-tocopherol methyltransferase ($\gamma$-TMT/VTE4) from perilla under the control of the cauliflower mosaic virus (CaMV) 35S promoter. The transgenic tobacco was confirmed by PCR and RT-PCR. The total content and composition of tocopherols in the transgenic lines were similar with wild type controls. However, chlorophyll-a and carotenoid content in the transgenic lines were increased by up to 45% (P<0.01) and 39% (P<0.02), respectively. Also, the over-expression of $\gamma$-TMT increased the salt stress tolerance in tobacco plants. These results demonstrate that over-expression of $\gamma$-TMT gene in tocopherol bio-synthetic pathway can increase salt stress tolerance and contents of chlorophyll-a and carotenoid in transgenic tobacco plants.