• Journal of Plant Biotechnology
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• v.32 no.3
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• pp.161-165
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• 2005

Agrobacterium tumefaciens-mediated cotyledonary explants transformation was used to produce transgenic cucumber. Cotyledonary explants of cucumber (c.v., Eunchim) were co-cultivated with strains Agrobaderium (LBA4404, GV3101, EHA101) containing the binary vector (pPTN289) carrying with CaMV 355 promoter-gus gene as reporter and NOS promoter-bar gene conferring resistance to glufosinate (herbicide Basta) as selectable marker. There was a significant difference in the transformation frequency depending Agrobacterium strains. The EHA101 of bacterial strains employed gave the maximum frequency (0.35%) for cucumber transformation. Histochemical gus and leaf painting assay showed that 15 individual lines were transgenic with the gus and bar gene. Southern blot analysis also revealed that the gus gene was successfully integrated into each genome of transgenic cucumber.

• Journal of Plant Biotechnology
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• v.30 no.1
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• pp.97-102
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• 2003

Lettuce (Lactuca sativa) was transformed with Agrobacterium tumefacience LBA4404 containing human granulocyte macrophage colony stimulating factor (hGM-CSF) gene to produce in cell suspension cultures. Cell suspension culture was established using callus from transgenic lettuce plant. Integration of hGM-CSF gene into plant chromosome was confirmed through genomic PCR and Southern blot analysis. In addition, Northern blot analysis indicated the expression of the introduced hGM-CSF gene in transformed lettuce. The recombinant hGM-CSF was expressed in transgenic cell cultures derived from transgenic plants as a yield of about 149.0 $\mu\textrm{g}$/L in culture filtrate, which was determined by ELISA. These results demonstrated that transformed lettuce cell suspension cultures could be used as a production system of therapeutic proteins such as hGM-CSF.

• Korean Journal of Plant Resources
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• v.27 no.3
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• pp.242-248
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• 2014

This study was conducted to establish an efficient transformation system by using somatic embryogenesis in an important Korean native conifer, Korean fir (Abies koreana). Embryogenic masses were induced from mature zygotic embryos of the Korean fir on Schenk and Hildebrandt medium, which was supplemented with thidiazuron. For genetic transformation, the embryogenic masses were co-cultivated with a disarmed Agrobacterium tumefaciens strain C58/pMP90 containing the plasmid vector pBIV10 or LBA4404 containing the plasmid vector MP90. Both vectors contain the kanamycin resistance and beta-glucuronidase (GUS) reporter genes. A total of 48 lines of embryogenic masses were selected on mLV medium containing $50{\mu}g/mL$ of kanamycin after 4 weeks of culture, following 3 days of co-cultivation with A. tumefaciens strain C58/pMP90 carrying pBIV10 (none of the lines was cultivated with strain LBA4404 carrying MP90). Quantitative real-time PCR was performed, and high levels of GUS transcripts were observed in the 48 putative transgenic lines; however, the control (non-transgenic line) showed negative results. Results of histochemical staining showed that the expression of the GUS reporter gene was observed in somatic embryos that developed from the embryogenic masses of all 48 lines. Stably transformed cultures were successfully produced by co-cultivation with A. tumefaciens strain C58/pMP90 carrying pBIV10 in Korean fir. Here, we have reported an Agrobacterium-mediated gene transfer protocol via somatic embryogenesis that may be helpful in developing breeding and conservation strategies for the Korean fir.

• Journal of The Korean Society of Grassland and Forage Science
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• v.21 no.3
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• pp.151-156
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• 2001

This study was conducted to obtain the transformed alfalfa (Medicago sativa L.) plants with thermotolerance gene (BcHSP17.6) using Agrobacterium tumefaciens LBA4404 and we confirmed the transformed gene from the regenerated alfalfa plants. The expression vector, pBKH4, harboring BcHSP17.6 gene was used for production of transgenic alfalfa plants. In a process for transformation, the callus of alfalfa was cocultivated with Agrobacterium tumefaciens and transformed calli were selected on kanamycin-containing SH-3-kc medium to regenerate into into the plant. The complete transgenic alfalfa plants were produced by cultivation for about 4 months on several regeneration media, SH-nk-c, SH-l lb-c, SH-sp-c, and SH-IBA. The transgenic alfalfa plants were analyzed by isolation of genomic DNA and PCR/Southem blot.

• Korean Journal of Plant Tissue Culture
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• v.25 no.3
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• pp.201-206
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• 1998

To develop the fruits of cucumber (Cucumis sativus L.) producing high yields of superoxide dismutase (SOD), the MnSOD cDNA from pea (Pisum sativum) under the control of the cauliflower mosaic virus 35S promoter was introduced into cucumber using Agrobacterium tumefaciens (strain LBA 4404)-mediated transformation. The kanamycin-resistant shoots were selected on the selection medium containing MS basal salt, 1.0 mg/L zeatin, 0.1 mg/L IAA, 300 mg/L claforan, and 100 mg/L kanamycin. After 6 weeks of culture on the selection medium, the shoots were transferred to MS medium containing 0.2 mg/L NAA to induce roots. PCR analysis using the primers for neomycin phosphotransferase (NPTII) gene revealed that three plantlets were transformed. The fruits of one transgenic plant had approximately 3.2-fold higher SOD activity than those of non-transgenic plants. MnSOD isoenzyme band was strongly detected on native gel in fruits of transgenic plants.

• Korean Journal of Plant Tissue Culture
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• v.25 no.4
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• pp.225-229
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• 1998

Agrobacterium tumefaciens LBA 4404 harboring binary vector pBI 121 was used for genetic transformation of lettuce(Lactuca Sativa L.). Optimal shoot regeneration from cotyledon explants was obtained in MS medium supplemented with 0.1mg/L NAA and 1.0 mg/L 2ip. In this condition, cotyledon explants were cocultivated with A, tumefaciens for 2 days, and then transferred to selection medium supplemented with 50 mg/L kanamycin and 500 mg/L carbenicillin. These explants were subsequently subcultured every 2 weeks on shoot induction medium. PCR analysis indicated that the GUS gene was stably integrated into the nuclear genome of lettuce. Histochemical analysis based on the enzymatic activity of the CUS protein showed that GUS activity was associated with vascular tissue in leaves and roots. Progenies of Ro plants demonstrated a linked monogenic segregation for GUS gene.

• Korean Journal of Plant Tissue Culture
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• v.25 no.4
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• pp.267-271
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• 1998

Cotyledon explants of lettuce were cocultured with Agrobacterium tumefaciens LBA4404::pBKS-GR1 harboring glutathione reductase(GR) gene in MS medium supplemented with 0.1 mg/L NAA and 1.0 mg/L 2ip for 48 hr. These explants were transferred to MS medium supplemented with 0.1 mg/L NAA and 1.0 mg/L 2ip, 50 mg/L kanamycin, and 500 mg/L carbenicillin. After 4 weeks of subculture, kanamycin-resistant shoots were obtained on selection medium. Leaves of putative transformants survived on selection medium containing 100 mg/L kanamycin. Incoporation of the GR gene into lettuce was confirmed by PCR analysis of genomic DNA. Southern blot analysis showed that ECL-labeled GR gene was hybridized to the expected amplified genomic DNA fragment of about 1.8 kb from transgenic lettuce. The presence of mRNA in transgenic lettuce was confirmed by RT-PCR with total RNA of transgenic lettuce. In progeny test of transformants, R$_1$ seeds were resistant to kanamycin (200mg/L) on MS medium.

• Korean Journal of Plant Tissue Culture
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• v.28 no.2
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• pp.97-101
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• 2001

The transport and allocation of photosynthetic assimilate is an important regulatory factor in plant productivity, In order to modify assimilate partitioning in rice, transgenic plants containing a potato sucrose transporter (SuT) gene were developed. Calli derived from rice seeds (Oryza sativa L. cv Dongjin) were cocultured with A. tumefaciens LBA 4404 harboring the SuT gene. Calli were transferred to MS medium supplemented with 50 mg/L hygromycin, 500 mg/L carbenicillin, 2 mg/L kinetin, 0.1 mg/L NAA. After 2 weeks, hygromycin resistant shoots were obtained from the calli on the selection medium. Roots were induced from the putative transgenic shoots on rooting MS medium supplemented with 250 mg/L cabenicillin. Plant regeneration rate from the calli was about 150%. Stable incorporation of the potato SuT gene into rice genomic DNA was confirmed by PCR and Southern blot analysis.

• Journal of Life Science
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• v.12 no.1
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• pp.67-76
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• 2002

The effect of transgene inactivation in T2, T3 and F2 generations was analyzed in progeny seedlings which had been generated by Agrobacterium (LBA4404/pBI121)-mediated transformation in Arabidopsis thaliana. In a system which investigated in the expression of $\beta$-glucuronidase(GUS)gene in kanamycin-resistant (ke $n^{R}$)seedlings, GUS inactivated seedlings were observed in 5 of 12 tested lines of T2 generation and the frequency of GUS inactivation was approximately 2.3%. Lines with multi-copies of T-DNA exhibited severe GUS gene inactivation with the frequency of 5.8% in T2 generation. In T3 generation lines exhibited GUS gene inactivation with the frequency of 1.3%. In contrast, inactivation increased dramatically up to 12.6% in multi-copy T-DNA line. A similar phenomenon was also found in F2 progeny from a transgenic line which had been crossed with wild-type Arabidopsis plant, WS-O (GUS gene inactivation frequency 9.9%). These results indicate that the foreign gene introduced into the plant was inactivated progressively in its transmission during subsequent generations and the transgenic line with multi-copies of T-DNA tended to show more increased inactivation.

• The Plant Pathology Journal
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• v.39 no.3
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• pp.255-264
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• 2023

Sweet potato symptomless virus 1 (SPSMV-1) is a single-stranded circular DNA virus, belonging to the genus Mastrevirus (family Geminiviridae) that was first identified on sweet potato plants in South Korea in 2012. Although SPSMV-1 does not induce distinct symptoms in sweet potato plants, its co-infection with different sweet potato viruses is highly prevalent, and thus threatens sweet potato production in South Korea. In this study, the complete genome sequence of a Korean isolate of SPSMV-1 was obtained by Sanger sequencing of polymerase chain reaction (PCR) amplicons from sweet potato plants collected in the field (Suwon). An infectious clone of SPSMV-1 (1.1-mer) was constructed, cloned into the plant expression vector pCAMBIA1303, and agro-inoculated into Nicotiana benthamiana using three Agrobacterium tumefaciens strains (GV3101, LBA4404, and EHA105). Although no visual differences were observed between the mock and infected groups, SPSMV-1 accumulation was detected in the roots, stems, and newly produced leaves through PCR. The A. tumefaciens strain LBA4404 was the most effective at transferring the SPSMV-1 genome to N. benthamiana. We confirmed the viral replication in N. benthamiana samples through strand-specific amplification using virion-sense- and complementary-sense-specific primer sets.