• Molecules and Cells
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• v.22 no.1
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• pp.89-96
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• 2006

"Determinate" and "indeterminate" inflorescences in plants are controlled by a single recessive gene, for example, SELF-PRUNING (SP) in Solanum lycopersicum, TERMINAL FLOWER1 in Arabidopsis, CENTRORADIALIS in Antirrhinum, and CENTRORADIALIS-like gene in tobacco. Pepper (Capsicum annuum L.) is an indeterminate species in which shoots grow indefinitely. In this study, we cloned and characterized the pepper SP-like gene (CaSP). RT-PCR revealed that the CaSP transcript accumulates to higher levels in floral buds than in other organs. Comparison of genomic DNA and cDNA sequences from indeterminate and determinate pepper plants revealed the insertion of a single base in the first exon of CaSP in the determinate pepper plants. CaSP is annotated in linkage group 8 (chromosome 6) of the SNU2 pepper genetic map and showed similar synteny to SP in tomato. Transgenic tobacco plants overexpressing CaSP displayed late-flowering phenotypes similar to the phenotypes caused by overexpression of CaSP orthologs in other plants. Collectively, these results suggest that pepper CaSP is an ortholog of SP in tomato.

• Journal of The Korean Society of Grassland and Forage Science
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• v.25 no.4
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• pp.281-286
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• 2005

To develop transgenic birdsfoot trefoil (Lotus corniculatus L.) plants tolerant to environmental stress, Arabidopsis NDPK gene (AtNDPK) was introduced into birdsfoot trefoil plants using Agrobacterium-mediated transformation and expressed powerfully under the control of the SWPA2 promoter. The expression vector, pCAMBIA2300 was used for introduction of AtNDPK gene into birdsfoot trefoil plants. The transformed calli were selected on kanamycin containing medium and then regenerated. The transformed birdsfoot trefoil plants were cultivated fur 4 months on BOi2Y medium. Genomic DNA PCR and Southern blot analysis confirmed the incorporation of AtNDPK into the birdsfoot trefoil genome.

• Journal of Life Science
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• v.17 no.8 s.88
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• pp.1157-1165
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• 2007

VvMSA, a grapevine ASR which is highly inducible by sugar and abscisic acid signals was previously shown to be a transcription factor for a hexose transporter gene VvHT1. We isolated a cDNA clone, VlASR which is regulated temporally during the grape berry development by ACP RT-PCR (annealing control primer reverse transcriptase-polymerase chain reaction) and it proved identical to VvMSA. RT-PCR and real-time PCR analyses revealed that the VlASR gene was expressed in berries at fruit set and that its expression increased as berries aged but decreased at the late ripening stage. In order to understand the regulatory mechanism of the asr gene, a genomic fragment was cloned from grapevine. The genomic DNA was 1375 bp long and a sugar box (sucrose box 3 and sucrose responsive element 1) was identified in the 611 bp upstream region of the open reading frame. Analysis of the VlASR promoter::reporter gene fusion demonstrated that this promoter was expressed in transgenic Arabidopsis even without sucrose treatment. This result suggests that the ASR/VvHT1-mediated sugar/ABA signaling, previously reported in grapevine, may not function in Arabidopsis which has no ASR homologue.

• Journal of Life Science
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• v.23 no.1
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• pp.1-7
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• 2013

Understanding salt tolerance mechanisms is important for the increase of crop yields, and so, several screening approaches were developed to identify plant genes which are involved in salt tolerance of plants. Here, we transformed the Arabidopsis cDNA library into a salt-sensitive calcineurin (CaN)-deficient ($cnb{\Delta}$) yeast mutant and isolated the colonies which can suppress salt-sensitive phenotype of $cnb{\Delta}$ mutant. Through this functional complementation screen, a total of 34 colonies functionally suppressed the salt-sensitive phenotype of $cnb{\Delta}$ yeast cells, and sequencing analysis revealed that these are 9 genes, including CaS, AtSUMO1 and AtHB-12. Among these genes, the ectopic expression of CaS gene increased salt tolerance in yeast, and CaS transcript was up-regulated under high salinity conditions. CaS-antisense transgenic plants showed reduced root elongation under 100 mM NaCl treatment compared to the wild type plant, which survived under 150 mM NaCl treatment, whereas CaS-antisense transgenic plant leaves turned yellow under 150 mM NaCl treatment. These results indicate that the expression of CaS gene is important for stress tolerance in yeast and plants.

• Journal of Plant Biotechnology
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• v.2 no.1
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• pp.43-49
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• 2000

An efficient and reliable Agrobacterium transformation procedure based on TDZ (thidiazuron)-induced organogenesis was established and applied to six Brazilian eggp1ant varieties. Optimum transgenic plants recovery was achieved upon the study of the following parameters affecting transformation efficiency, using F-100 variety as a model: i) explant source; ii) pre-culture period; iii) physical state of the pre-culture medium and iv) coculture conditions. The highest frequency of kanamycin-resistant calli derived from leaf explants (5%) was obtained without a pre-culture period and co-cultivation for 24 h in liquid medium followed by five days on solid RM (regeneration medium). For cotyledon explants, best results were achieved upon a pre-culture of 24 h in liquid RM and a co-cultivation period of 24 h in liquid RM followed by three days in solid RM, resulting in a transformation Sequency of 22.7%. Kanamycin-resistant organogenic calli were also obtained from cultivars Emb, Preta Comprida, Round nose Shaded, Campineira and Florida Market. The expression pattern of an epidermis-specific promoter was studied using transformants expressing a chimaeric construct comprised by the promoter Atgrp-5 transcriptionally fused to the coding region of the gus gene. The expression pattern was similar to that previously observed in tobacco and Arabidopsis thaliana, with preferential expression at the epidermis and the stem phloem. These results support the idea that the Atgrp-5 promoter can be used to drive defense genes in these tissues, which are sites of pathogen interaction and spread, in programs for the genetic improvement of eggplant.

• The Plant Pathology Journal
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• v.23 no.2
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• pp.76-89
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• 2007

CaCDPK4, a full-length cDNA clone encoding Capsicum annuum calcium-dependent protein kinase 4, was isolated from chili pepper (Capsicum annuum L.). Deduced amino acid sequence of CaCDPK4 shares the highest homology with tobacco NpCDPK8 and chickpea CaCDPK2 with 79% identity. Genomic blot analyses revealed that CaCDPK4 is present as a single copy in pepper genome, but it belongs to a multigene family. CaCDPK4 was highly induced when pepper plants were inoculated with an incompatible bacterial pathogen. Induced levels of CaCDPK4 transcripts were also detected in pepper leaves by the treatment of ethephon, an ethylene-inducing agent, and high-salt stress condition. The bacterial-expressed GST-CaCDPK4 protein showed to retain the autophosphorylation activity in vitro. GUS expression driven by CaCDPK4 promoter was examined in transgenic Arabidopsis containing transcriptional fusion of CaCDPK4 promoter. GUS expression under CaCDPK4 promoter was strong in the root and veins of the seedlings. GW (-1965) and D3 (-1377) promoters conferred on GUS expression in response to inoculation of an incompatible bacterial pathogen, but D4-GUS (-913) and DS-GUS (-833) did not. Taken together, our results suggest that CaCDPK4 can be implicated on signal transduction pathway of defense response against an incompatible bacterial pathogen in pepper.

• Korean Journal of Plant Tissue Culture
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• v.27 no.5
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• pp.407-409
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• 2000

Outer envelope membrane proteins of chloroplasts encoded by the nuclear genome are transported without the N-terminal transit peptide. Here, we investigated the targeting mechanism of AtOEP7, an Arabidopsis homolog of small outer envelope membrane proteins in vivo. AtOEP7 was expressed transiently in protoplasts or stably in transgenic plants as fusion proteins with GFP. In both cases AtOEP7:GFP was targeted to the outer envelope membrane when assayed under a fluorescent microscope or by Western blot analysis. Except the transmembrane domain, deletions of the N- or C-terminal regions of AtOEP7 did not affect targeting although a region closed to the C-terminal side of the transmembrane domain affected the targeting efficiency. Targeting experiments with various hybrid transmembrane mutants revealed that the amino acid sequence of the transmembrane domain determines the targeting specificity The targeting mechanism was further studied using a fusion protein, AtOEP7：NLS：GFP, that had a nuclear localization signal. AtOEP7：NLS：GFP was efficiently targeted to the chloroplast envelope despite the presence of the nuclear localization signal. Taken together, these results suggest that the transmembrane domain of AtOEP7 functions as the sole determinant of targeting specificity and that AtOEP7 may be associated with a cytosolic component during translocation to the chloroplast envelope membrane.

• Molecules and Cells
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• v.40 no.11
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• pp.828-836
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• 2017

Eukaryotic cells consist of a complex network of thousands of proteins present in different organelles where organelle-specific cellular processes occur. Identification of the subcellular localization of a protein is important for understanding its potential biochemical functions. In the post-genomic era, localization of unknown proteins is achieved using multiple tools including a fluorescent-tagged protein approach. Several fluorescent-tagged protein organelle markers have been introduced into dicot plants, but its use is still limited in monocot plants. Here, we generated a set of multicolored organelle markers (fluorescent-tagged proteins) based on well-established targeting sequences. We used a series of pGWBs binary vectors to ameliorate localization and co-localization experiments using monocot plants. We constructed different fluorescent-tagged markers to visualize rice cell organelles, i.e., nucleus, plastids, mitochondria, peroxisomes, golgi body, endoplasmic reticulum, plasma membrane, and tonoplast, with four different fluorescent proteins (FPs) (G3GFP, mRFP, YFP, and CFP). Visualization of FP-tagged markers in their respective compartments has been reported for dicot and monocot plants. The comparative localization of the nucleus marker with a nucleus localizing sequence, and the similar, characteristic morphology of mCherry-tagged Arabidopsis organelle markers and our generated organelle markers in onion cells, provide further evidence for the correct subcellular localization of the Oryza sativa (rice) organelle marker. The set of eight different rice organelle markers with four different FPs provides a valuable resource for determining the subcellular localization of newly identified proteins, conducting co-localization assays, and generating stable transgenic localization in monocot plants.

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2005.11a
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• pp.9-21
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• 2005

To study the biochemical and physiological role of the plastidic glucose transporter (pGlcT) in carbohydrate metabolism, we characterized transgenic plants with mutations in the pGlcT gene (GT), gt-1 and gt-2, as well double mutants of GT and the maltose transporter (MEX1) and GT and the triose phosphate/phosphate translocator (TPT), GT and the cytosolic fructose-1,6-bisphosphatase gene (cFBP), and MEX1 and TPT, gt-1/mex2, gt-1/tpt-2, gt-1/cfbp-1, mex1-1/tpt-2, respectively. Compared to the wild type, all mutants except the gt-1/cfbp-1 mutant lines displayed higher starch accumulation and higher levels of maltose. Starch accumulation is due to a decrease in starch turnover, leading to an imbalance between the rates of synthesis and degradation. Sucrose levels of gt alleles were higher than those in wild-type plants during the light period, suggesting possible nightly supplementation via the maltose transport pathway to maintain proper carbohydrate partitioning in the plant leaves. The gt plants displayed less growth retardation than mex1-1 mutant and gt-1/mex2 double mutant displayed accumulativesevere growth retardation as compared to individual gt-1 and mex1-1 mutants, implying that the maltose transporter-mediated pathway is a major route for carbohydrate partitioning at night. The gt-1/tpt-2, mex1-1/tpt-2 and gt-1/cfbp-1 double mutants had retarded growth and low chlorophyll content to differing degrees, indicating that photosynthetic capacity had diminished. Interestingly, the gt-1/tpt-2 line displayed a glucose-insensitive phenotype and higher germination rates than wild type, suggesting its involvement not only in carbon partitioning, but also in the sugar signaling network of the pGlcT and TPT.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.54 no.4
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• pp.375-383
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• 2009

This study was carried out to develop transgenic rice cultivars with the CAX1 (accession no. U57411) gene. We successfully selected the transgenic rice plants over-expressing the Arabidopsis H+/$Ca^{2+}$ antiporter CAX1 (accession no. U57411) gene in T6 generation. The brown rice of the CAX1 expressing rice contained 13.4~68.0 % more calcium $(Ca^{2+})$ than that of the wild type and 5 lines were selected based on the phenotypes compared to the control cultivar at the GMO field. The CAX1 expressing transgenic rice plants were similar in phenotype to the wild type during the whole growth period. Also these selected 4 lines appeared to be resistant to blast, cold and water solution compared with the wild type. Difference in 1,000 grain weight of brown rice was observed among each line but grain shape did not show any morphological alternations. These results suggest the enhanced Ca-substrate specificity of CAX1 exchanger in donor plant. Therefore, intact CAX1 exchanger can be functionally useful for $Ca^{2+}$ nutrient enrichment of rice with reduced accumulation of undesirable cation.