• 한국연초학회지
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• 제20권1호
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• pp.66-70
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• 1998

TMV resistant lines (TRLs) originated from the Blo plant of Nicotiana tabacum cv. NC82 transformed with TMV coat protein cDNA which initially showed delayed disease symptom were selected for increased resistance in each subsequent generation. The result of field experiment of the transgenic tobacco lines in the fifth generation for TMV resistance and their response to other tobacco diseases (black shank, bacterial wilt, and powdery mildew) is described in this report. When fifteen TRLs of the fifth generation were tested for TMV resistance by mechanically inoculating the individual plants, over 95 percent of the plants of 6 lines showed complete resistance even 8 weeks after the inoculation. Average frequency of the resistant plants in TRLs of the fifth generation 8 weeks after the inoculation was 87%. Stable insertion and expression of TMV coat protein cDNA in the fifth generation of the transgenic tobacco plant were confirmed by PCR and immunoblot hybridization, respectively. All TRLs were resistant to the black shank but were susceptible to the bacterial wilt disease and the powdery mildew to the same degree as non-transgenic NC82 was. Therefore, it was indicated that the phenotypes related at least to disease resistance were not changed in the transgenic tobacco. Key words : TMV CP cDNA, TMV resistant tobacco plant, transformation.

• Plant Biotechnology Reports
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• 제4권1호
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• pp.75-83
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• 2010

Glycine betaine has been reported as an osmoprotectant compound conferring tolerance to salinity and osmotic stresses in plants. We previously found that the expression of betaine aldehyde dehydrogenase 1 gene (OsBADH1), encoding a key enzyme for glycine betaine biosynthesis pathway, showed close correlation with salt tolerance of rice. In this study, the expression of the OsBADH1 gene in transgenic tobacco was investigated in response to salt stress using a transgenic approach. Transgenic tobacco plants expressing the OsBADH1 gene were generated under the control of a promoter from the maize ubiquitin gene. Three homozygous lines of $T_2$ progenies with single transgene insert were chosen for gene expression analysis. RT-PCR and western blot analysis results indicated that the OsBADH1 gene was effectively expressed in transgenic tobacco leading to the accumulation of glycine betaine. Transgenic lines demonstrated normal seed germination and morphology, and normal growth rates of seedlings under salt stress conditions. These results suggest that the OsBADH1 gene could be an excellent candidate for producing plants with osmotic stress tolerance.

• 식물조직배양학회지
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• 제28권1호
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• pp.41-45
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• 2001

CAB 유전자로 형질전환된 담배 2세대 식물체의 도입된 CAB 유전자 존재여부를 genomic PCR 방법으로 각계통에서 확인하였다. CAB유전자로 형질전환된 2세대 식물체를 자연 광 조건과 90% 차광된 온실에서 각각 생육시킨 결과 형질전환 식물체의 광합성능 정도는 정상 식물체와 유사하거나 약간 높은 경향이었으며, 광포화점은 형질전환 담배 식물체나 정상 식물체 모두 500$\mu$mol m$^{-2}$ s$^{-1}$로 차이가 없었다. 조사한 7계통 중 C7, C11, C14 계통의 광합성 정도가 조사된 모든 광량에서 정상 담배 식물체보다 높게 나타났다. 차광된 조건에서 생육한 담배 식물체의 광합성능 정도는 조사된 7계통 중 C2, C11, C14 계통이 90% 차광된 온실조건에서도 광합성 능이 우수하게 나타났다. 형질전환 담배 식물체의 chlorophyll 함량은 정상 NC82와 차이가 없었으며, 양지에서 생육한 조건과 90% 차광된 조건 모두에서 차이점이 없었으며 광합성능과 chlorophyll 함량과의 유의성은 없었다. 형질전환 담배 식물체 수확엽의 건물률은 정상 식물체와 비교하여 차이점이 나타나지 않았으며, 내용성분도 nicotine, 전당, 전질소에서 차이가 없었다.

• Journal of Plant Biotechnology
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• 제2권1호
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• pp.25-28
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• 2000

The anthocyanin gene encoding flavonoid 3',5'-hydroxylase(F3,5H) was normally expressed in Nicotiana tobacco (Xanthi) plants cocultivated with Agrobacterium tumefaciens LBA4404 carrying egg plant flavonoid 3',5'-hydroxylase cDNA. Northern blot analysis showed the normal expression of F3', 5'H gene from transgenic plants. Here we found the phenotypic differences between transgenic plants and wild-type plants. The petal shape of transgenic plants showed more round shape and around petal tube area was compared to that of wild-type tobacco plants. And the petal color of transgenic plants was much lighter than that of wild-type tobacco plants.

• 한국환경농학회지
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• 제27권4호
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• pp.389-394
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• 2008

The ecophysiological changes occurring upon cold stress were studied using cold tolerant transgenic and wild-type tobacco plants. In a previous study, cold tolerance in tobacco was induced by the introduction of a gene encoding the zinc finger transcription factor, PIF1. Gas-exchange measurements including net photosynthesis and stomatal conductance were performed prior to, in the middle of, and after a cold-stress treatment of $1{\pm}2^{\circ}C$ for 96 h in each of the four seasons. In both transgenic and wild-type plants, gas-exchange parameters were severely decreased in the middle of the cold treatment, but had recovered after 2-3 h of adaptation in a greenhouse. Most t-test comparisons on gas-exchange measurements between the two plant types did not show statistical significance. Wild-type plants had slightly more water-soaked damage on the leaves than the transgenic plants. A light-response curve did not show any differences between the two plant types. However, the curve for assimilation-internal $CO_2$ in wild-type plants showed a much higher slope than that of the PIF1 transgenic plants. This means that the wild-type plant is more capable of regenerating Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and has greater electron transport capacity. In conclusion, cold-resistant transgenic tobacco plants demonstrated a better recovery of net photosynthesis and stomatal conductance after cold-stress treatment compared to wild-type plants, but the ecophysiological recoveries of the transgenic plants were not statistically significant.

• Journal of Plant Biotechnology
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• 제4권4호
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• pp.143-150
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• 2002

A strong oxidative stress-inducible peroxidase promoter (referred to as SWPA2 promoter) was cloned from tell cultures of sweetpotato (Ipomoea batatas) and characterized in transgenic tobacco cultured cells in terms of biotechnological applications. Employing a transient expression assay in tobacco protoplasts, with five different 5'-deletion mutants of the SWPA2 promoter fused to the $\beta$-glucuronidase (GUS) reporter gene, the 1314 bp deletion mutant showed approximately 30 times higher GUS expression than the CaMV 35S promoter. The expression of GUS activity in suspension cultures of transgenic cells derived from transgenic tobacco leaves containing the -1314 bp SWPA2 promoter-GUS fusion was strongly expressed following 15 days of subculture compared to other deletion mutants, suggesting that the 1314 bp SWPA2 promoter will be biotechnologically useful for the development of transgenic cell lines engineered to produce key pharmaceutical proteins. In this respect, we developed transgenic cell lines such as tobacco (Nicotiana tabacum L. BY-2), ginseng (Panax ginseng) and Siberian ginseng (Acanthopanax senticosus) using a SWPA2 promoter to produce a human lactoferrin (hLf) and characterized the hLf production in cultured cells. The hLf production monitored by ELISA analysis in transgenic BY-2 cells was directly increased proportional to cell growth and reached a maximal level (up to 4.3% of total soluble protein) at the stationary phase in suspension cultures. The SWPA2 promoter should result in higher productivity and increased applications of plant cultured cells for the production of high-value recombinant proteins.

• BMB Reports
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• 제34권4호
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• pp.316-321
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• 2001

Dehydroascorbate (DHA) reductase (DHAR, EC 1.8.5.1) catalyzes the reduction of DHA to reduced ascorbate (AsA) using glutathione (GSH) as the electron donor in order to maintain an appropriate level of ascorbate in plant cells. To analyze the physiological role of DHAR in environmental stress adaptation, we developed transgenic tobacco (Nicotiana tabacum cv. Xanthi) plants that express a human DHAR gene isolated from the human fetal liver cDNA library in the chloroplasts. We also investigated the DHAR activity, levels of ascorbate, and GSH. Two transgenic plants were successfully developed by Agrobacterium-mediated transformation and were confirmed by PCR and Southern blot analysis. DHAR activity and AsA content in mature leaves of transgenic plants were approximately 1.41 and 1.95 times higher than in the non-transgenic (NT) plants, respectively In addition, the content of oxidized glutathione (GSSG) in transgenic plants was approximately 2.95 times higher than in the NT plants. The ratios of AsA to DHA and GSSG to GSH were changed by overexpression of DHAR, as expected, even though the total content of ascorbate and glutathione was not significantly changed. When tobacco leaf discs were subjected to methyl viologen at $5\;{\mu}M$, $T_0$ transgenic plants showed about a 50% reduction in membrane damage compared to the NT plants.

• Journal of Microbiology and Biotechnology
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• 제20권5호
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• pp.917-924
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• 2010

The practicability of transgenic tobacco engineered to express bacterial native mercuric reductase (MerA), responsible for the transport of $Hg^{2+}$ ions into the cell and their reduction to elemental mercury ($Hg^0$), without any codon modification, for phytoremediation of mercury pollution was evaluated. Transgenic tobacco plants reduce mercury ions to the metallic form; take up metallic mercury through their roots; and evolve the less toxic elemental mercury. Transformed tobacco produced a large amount of merA protein in leaves and showed a relatively higher resistance phenotype to $HgCl_2$ than wild type. Results suggest that the integrated merA gene, encoding mercuric reductase, a key enzyme of the bacterial mer operon, was stably integrated into the tobacco genome and translated to active MerA, which catalyzes the bioconversion of toxic $Hg^{2+}$ to the least toxic elemental $Hg^0$, and suggest that MerA is capable of reducing the $Hg^{2+}$, probably via NADPH as an electron donor. The transgenic tobacco expressing merA volatilized significantly more mercury than wild-type plants. This is first time we are reporting the expression of a bacterial native merA gene via the nuclear genome of Nicotiana tabacum, and enhanced mercury volatilization from tobacco transgenics. The study clearly indicates that transgenic tobacco plants are reasonable candidates for the remediation of mercurycontaminated areas.

• 식물조직배양학회지
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• 제25권4호
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• pp.245-250
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• 1998

Tobacco mosaic virus(TMV) 외피단백질 유전자를 연초(Nicotiana tabacm cv. NC82)에 형질전환하고 형질전환 식물체 후세대에서 TMV 저항성인 연초를 선발하여, 선발된 TMV 저항성 제5세대 형질전환 식물체의 도입된 유전자발현 및 고온에서의 특성 등을 조사하였다. TMV 저항성 식물체의 염색체 DNA에 TMV 외피 단백질 유전자가 안정되게 존재하고 있음을 genomic PCR을 수행하여 확인하였다. 또한 형질전환 식물체내에서 TMV 외피 단백질 발현은 Immunoblot hybridization 방법으로 확인하였다. TMV 저항성 형질전환 연초식물체에서 발현된 단백질의 양은 매우 적었으며 특히 본엽에는 병징이 나타나지 않았으나 수확기 마지막 액아에 TMV의 반점이 나타난 병징발현 지연형의 형질전환 식물체의 경우에도 발현된 단백질의 양은 정상 NC 82에 TMV가 감염되었을 때와 비교하여 현저히 적었다. TMV 저항성 형질전환 식물체 내에서 발현되는 TMV 외피단백질의 양은 총 단백질에 대비하여 0.01% 이하이였다. TMV 병징 발현 지연형인 형질전환 식물체에 TMV를 인공접종한 후 고온처리상태에서 외피 단백질 유전자의 전사 및 발현을 RT-PCR과 Immune blot hybridization 통하여 확인하였으며, 이때 TMV의 증식도 억제되었으므로 개량멀칭시 나타나는 고온조건하에서도 저항성이 안정적으로 발현될 수 있음을 알 수 있었다.

• Plant Biotechnology Reports
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• 제3권2호
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• pp.157-165
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• 2009

Human erythropoietin (EPO) is a leading product in the biopharmaceutical market, but functional EPO has only been produced in mammalian cells, which limits its application and drives up the production costs. Using plants to produce human proteins may be an alternative way to reduce the cost. However, a recent report demonstrated that overexpression of the human EPO gene (EPO) in tobacco or Arabidopsis rendered males sterile and retarded vegetative growth, which raises concern whether EPO might interfere with hormone levels in transgenic plants. In the present study, we demonstrated that overexpressing EPO with additional 5'-His tag and 3' ER-retention peptides in tobacco did not cause any developmental defect compared to GUS plants. With our method, all 20 transgenic plants grew on selective medium and, further confirmed by PCR, were fertile. Most of them grew similarly compared to GUS plants. Only one transgenic plant (EPO2) was shorter in plant height but had twice the life span compared to other transgenic plants. When 11 randomly selected EPO plants, along with the abnormal plant EPO2, were subjected to RT-PCR analysis, all of them had detectable EPO transcripts. However, their protein levels varied considerably; seven of them had detectable EPO proteins analyzed by western blot. Our results indicate that overexpressing human EPO protein in plants does not have detrimental effects on growth and development. Our transformation systems allow us to further explore the possibility of glycoengineering tobacco plants for producing functional EPO and its derivatives.