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Cloning of Molecular Marker for Cultivar Protection and Transfer to Nicotiana tabacum L. (품종보호를 위한 분자 마커의 Cloning 및 담배로의 전이)

  • Ku, Ja Jung;Park, Young Doo;Choi, Geun Won
    • Horticultural Science & Technology
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    • v.17 no.6
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    • pp.770-772
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    • 1999
  • This study was conducted to provide a basic system to develop a molecular marker for plant cultivar protection using a recombinant DNA technology. Using Nicotiana tabacum L. plants, the potentiality in the utilization of the developed marker was examined. After homology test with several plant genomes, mouse adenosine deaminase (ADA) gene was selected as DNA source of a molecular marker for cultivar protection. As a result of the digestion of ADA gene with BamHI and Pst I, six DNA fragments were obtained, and 513 bp DNA fragment among them was selected as a possible DNA marker for cultivar protection. Selected 513 bp DNA fragment was efficiently inserted into pBI101 plasmid vector for plant transformation by using phagemid vector pBluescript II SK (+/-) as an intermediate vector. The recombinant pBI101, carrying 513 bp DNA fragment, possible markers for cultivar protection, was transformed into A. tumefaciens LBA4404. Nicotiana tabacum was transformed with A. tumefaciens LBA4404 having the recombinant pBI101 and was confirmed the transfer of 513 bp DNA fragment, a possible molecular marker for cultivar protection.

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Loss of Function in GIGANTEA Gene is Involved in Brassinosteroid Signaling

  • Hwang, Indeok;Park, Jaeyoung;Lee, Beomgi;Cheong, Hyeonsook
    • Journal of Integrative Natural Science
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    • v.4 no.2
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    • pp.113-120
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    • 2011
  • Brassinosteroids (BRs) are plant steroid hormones that play essential roles in growth and development. Mutations in BR-signaling pathways cause defective in growth and development like dwarfism, male sterility, abnormal vascular development and photomorphogenesis. Transition from vegetative to reproductive growth is a critical phase change in the development of a flowering plant. In a screen of activation-tagged Arabidopsis, we identified a mutant named abz126 that displayed longer hypocotyls when grown in the dark on MS media containing brassinazole (Brz), an inhibitor of BRs biosynthesis. We have cloned the mutant locus using adapter ligation PCR walking and identified that a single T-DNA had been integrated into the ninth exon of the GIGANTEA (GI) gene, involved in controling flowering time. This insertion resulted in loss-of-function of the GI gene and caused the following phenotypes: long petioles, tall plant height, many rosette leaves and late flowering. RT-PCR assays on abz126 mutant showed that the T-DNA insertion in GIGANTEA led to the loss of mRNA expression of the GI gene. In the hormone dose response assay, abz126 mutant showed: 1) an insensitivity to paclobutrazole (PAC), 2) an altered response with 6-benzylaminopurine (BAP) and 3) insensitive to Brassinolide (BL). Based on these results, we propose that the late flowering and tall phenotypes displayed by the abz126 mutant are caused by a loss-of-function of the GI gene associated with brassinosteroid hormone signaling.

Evidence for Existence of a Water-Extractable Anticoagulant in an Earthworm, Lumbricus rubellus

  • Woo, Jeong-Im;Bahk, Yun-Kyung;Yu, Kyoung-Hee;Paik, Seung-R.;Chang, Chung-Soon
    • BMB Reports
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    • v.29 no.6
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    • pp.500-506
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    • 1996
  • We have isolated a water-extracted novel regulator for blood coagulation from an earthworm, Lumbricus rubellus. As a folk remedy, the earthworm has been known to facilitate blood circulation. After complete heat inactivation of endogenous proteases in the earthworm, an anticoagulant(s) was purified through ammonium sulfate fractionation and three consecutive gel permeation chromatography of Sephacryl S-300, Sephadex G-75, and G-150 by measuring activated partial thromboplastin time (APTT) The anticoagulant was further purified to 2,800 fold with a C4 reversed-phase HPLC This activity was stable under heat ($100^{\circ}C$ for 30 min) and acidic conditions (0.4 N HCl). The effects of this partially purified anticoagulant on thrombin were observed with various substrates such as N${\alpha}$-benzoyl-DL-arginine-p-nitroanilide (BApNA), H-D-phenylalanyl-L-pipecoyl-L-arginine-p-nitroanilide (S-2238), N${\alpha}$-p-tosyl-L-arginine methyl ester (TAME), and fibrinogen as a natural substrate. Only TAME hydrolysis, due to an esterase activity of the enzyme, was inhibited among the chromogenic substrates. In addition, the anticoagulant not only inhibited the conversion of fibrinogen to fibrin but also prolonged the fibrin clot formation monitored with the in vitro coagulation test. Based on these observations, we suggest the significance of measuring the ability of antithrombotic drugs to inhibit the esterase activity of thrombin. In this report, it was also shown that the earthworm indeed contained a water-extractable, heat- and acid-stable anticoagulant which could be used as a novel antithrombotic agent.

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Distinctive response of maize (Zea mays L.) genotypes in vitro with the acceleration of phytohormones

  • Muppala, Sridevi;Gudlavalleti, Pavan Kumar;Pagidoju, Sreenu;Malireddy, Kodandarami Reddy;Puligandla, Sateesh Kumar;Dasari, Premalatha
    • Journal of Plant Biotechnology
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    • v.47 no.1
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    • pp.26-39
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    • 2020
  • In maize, immature embryos (IEs) are highly regenerative explants most suitable for producing high frequencies of plantlet regeneration in vitro. Apart from media, explants, and hormones, genotypic variation also influences in vitro characters to a great extent. In the present study, IEs were used to study the distinctive effect of variation of size/stage and hormones in different genotypes on five in vitro characters viz., frequency of callus induction, growth rate of total callus, frequency of E. callus induction, and volume and number of regenerated plantlets. LS medium with different concentrations of 2,4-D (0.5, 1.5, 2.5, 4.0 and 5.0 mg/L) were used to study the former four in vitro characters, and medium with 6-benzylaminopurine and kinetin (0.5 mg/L, each) was used for plantlet regeneration. IEs of 1.0, 1.5, 2.0, 2.5 and 3.0 mm in size were isolated from four inbred lines viz., NM74C, NM81A, NM5883 and NM5884. Two-way ANOVA revealed that explant size and genotypes, as well as hormonal concentrations showed significant effects on in vitro characters. Two millimeter IEs were found to be suitable for in vitro cultures. LS medium with 1.5 mg/L 2,4-D and LS with BAP and Kn (0.5 mg/L, each) were found to be the best hormonal concentrations for callus induction, maintenance, and regeneration, respectively. Among the four genotypes, NM81A and NM5883 yielded more non-embryogenic and Type I E. calli. In contrast, NM74C and NM5884 yielded more highly regenerative Type II calli. Inbred line NM5884 was found to be the best among these four genotypes.

Tissue-cultured regeneration and ecological values in major bamboo species

  • Sharma, Avinash;Manpoong, Chowlani;Gohain, Anwesha;Pandey, Himanshu;Padu, Gompi;Aku, Hage
    • Journal of Ecology and Environment
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    • v.46 no.3
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    • pp.218-242
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    • 2022
  • Background: Promising specific growth regulators are employed in the tissue cultures of various bamboo species. Specific natural hardening mixtures support the acclimatization and adaptation of bamboo under protected cultivation. Results: The growth regulators like 2, 4-Dichlorophenoxyacetic acid (2, 4-D), Naphthaleneacetic Acid (NAA), Thidiazuron (TDZ), 6-Benzylaminopurine (BAP), Kinetin, Gelrite, Benzyl Adenine (BA), Indole Butyric Acid (IBA), Coumarin, Putrescine, Gibberellic acid (GA3), Indole Acetic Acid (IAA) has been widely used for callus induction, root regeneration and imposing plant regeneration in various species of bamboo such as Bambusa spp. and Dendrocalamus spp. Different combinations of growth regulators and phytohormones have been used for regenerating some of the major bamboo species. Natural hardening materials such as cocopeat, vermicompost, perlite, cow dung, farmyard manure, compost, soil, garden soil, and humus soil have been recommended for the acclimatization and adaptation of bamboo species. Standard combinations of growth regulators and hardening mixtures have imposed tissue culture, acclimatization, and adaptation in major bamboo species. Conclusions: Bamboo contributes to soil fertility improvement and stabilization of the environment. Bamboo species are also involved in managing the biogeochemical cycle and have immense potential for carbon sequestration and human use. This paper aims to review the various growth regulators, natural mixtures, and defined media involved in regenerating major bamboo species through in vitro propagation. In addition, the ecological benefits of safeguarding the environment are also briefly discussed.

Development of transgenic disease-resistance root stock for growth of watermelon.(oral)

  • S.M. Cho;Kim, J.Y.;J.E. Jung;S.J. Mun;S.J. Jung;Kim, K.S.;Kim, Y.C.;B.H. Cho
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.65.2-65
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    • 2003
  • To protect the plant against several soil-borne pathogens, we are currently constructing disease-resistant transgenic root stock for the growth of cucurbitaceae vegetable plants, watermelon and gourd. We made a watermelon cDNA library from Cladosporium cucumerinum-Infected leaves for substractive hybriazation and differential screening. We isolated the several pathogen inducible cDNA clones, such as caffeoyl-CoA-methyltransferase, LAA induced protein, receptor-like kinase homolog, hydroxyproline-rich glycoprotein, catalase, calmodulin binding protein, mitochondrial ATPase beta subunit, methyl tRNA synthetase and WRKY transcription factors. We previously obtained CaMADS in pepper and galactinol synthase ( CsGolS) in cucumber that were confirmed to be related with disease-resistance. CaMADS and CsGolS2 were transformed into the inbred line 'GO701-2' gourd, the inbred line '6-2-2' watermelon and the Kong-dye watermelon by Agrobacterium tumerfaciens LBA4404. Plant growth regulators (zeatin, BAP and IAA) were used for shoot regeneration and root induction for optimal condition. Putative transgenic plants were selected in medium containing 100mg/L kanamycin and integration of the CaMADS and CsGO/S2 into the genomic DNA were demonstrated by the PCR analysis. We isolated major soil-borne pathogens, such as Monosporascus cannonballus, Didymella bryoniae, Cladosporium cuvumerinum from the cultivation area of watermelon or root stock, and successfully established artificial inoculation method for each pathogen. This work was supported by a grant from BioGreen 21 program, Rural Development Administration, Republic of Korea.

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Randomized, Double-blind, and Placebo-controlled a Clinical Study for Chronic Fatigue via the Analysis of Efficacy and Safety of Gongjin-dan and Ssanghwa-tang : Study Protocol (만성피로에 대한 공진단과 쌍화탕의 안전성 및 유효성 평가를 위한 무작위배정, 이중눈가림, 위약대조 임상시험 : 임상연구 프로토콜)

  • Jun-Yong, Choi;Byungmook, Lim;Hyeun-kyoo, Shin;Kibong, Kim
    • Journal of Society of Preventive Korean Medicine
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    • v.26 no.3
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    • pp.97-108
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    • 2022
  • Objectives : The purpose of this study is to confirm the efficacy and safety of Gongjin-dan and Ssanghwatang for chronic fatigue. Methods : A total of 90 people, between 19 and 65 years old, will be recruited to participate in a randomized, double-blind, and placebo-controlled a clinical trial. Participants in the Gongjin-dan group will take one pill of Gongjin-dan along with three packs of placebo oral liquid Ssanghwa-tang per day for 4 weeks. Participants in the Ssanghwa-tang group will take three packages of liquid Ssanghwa-tang and one placebo Gongjindan pill per day for 4 weeks. In the placebo group, participants will take one pill of placebo Gongjin-dan and three packs of placebo liquid Ssanghwa-tang per day, for 4 weeks. Outcomes will be measured at the baseline, 4th week, and 6th week. The primary outcome is the change in the Fatigue Severity Scale (FSS). Secondary outcomes are the change of Multidimensional Fatigue Inventory-20 (MFI-20), Chalder Fatigue Scale (CFQ), Short-Form 36 Health Survey (SF-36), Korean Version of Schedule of Fatigue and Anergy/General Physician (SOFA/GP), Glucose, Lactate, Ammonia, Free Fatty Acid (FAA), d-ROMs&BAP, Selenium, and Cortisol. Results : This trial was approved by the institutional review board of Pusan National University Korean Medicine Hospital (registry number: PNUKHIRB 2021-10-005). Recruitment opened in November 2021 and is supposed to be completed by December 2022. Conclusions : This trial will provide clinical information to determine the efficacy and safety of Gongjindan and Ssanghwa-tang for chronic fatigue.

Cell Growth in Suspension-Culture of Populus nigra var. italica and the Efficiency of Micro-Callus Formation according to Cell Plating Method (Populus nigra var. italica현탁배양(懸濁培養) 세포(細胞)의 생장(生長) 및 Cell Plating방법(方法)에 따른 Micro-Callus형성능력(形成能力))

  • Kim, Chi Moon;Lee, Jae Soon;Kwon, Ki Won
    • Korean Journal of Agricultural Science
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    • v.14 no.2
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    • pp.197-204
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    • 1987
  • In order to know the growth of suspended cells by explant sources, the change of nitrogen contents of cultured cells following the growth periods, capability of micro-callus formation according to cell plating methods, growth of suspended cells on various media, and efficiency of micro-callus formation by using growth regulators and different N strengths were investigated. 1. When suspension culture was tried by using the callus induced from internode and petiole, cell fresh weight and packed cell volume increased with similar way and the growth reached at stationary phase after 12 culture days. 2. N-contents of cultured cells increased upto 3 days and decreased around 6days. But the values increased again upto 9 days, after that they showed gradual decreases. 3. Of cell plating methods, embedding method was the best for micro-callus formation. 4. Growth of suspened cells showed the rest performanoes, when they were cultured on LM medium with 1/2N strengths and BAP 0.01.2.4-D 0.1, and NAA $1.0mg/{\ell}$, after 15 cultured days(upto 76.9 folds). LM medium was better than MS or GD. The combination of auxin and cytokinin was better for cell growing than auxin-treatment only. 5. Micro-callus from single cell and small cell aggregates was formed only on MS and LM media with 2,4-D $1.0mg/{\ell}$.

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Effects of l-arginine supplementation with high-intensity training on muscle damage and fatigue index and athletic performance in Canoe Athletes (L-arginine 섭취가 고강도 훈련 프로그램에 따른 카누선수의 근 손상 지표, 피로 물질 및 경기력 향상에 미치는 영향)

  • Jung, Jong-Hwan;Kang, Eun-Bum;Kim, Chang-Hwan
    • Journal of the Korean Applied Science and Technology
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    • v.36 no.3
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    • pp.942-953
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    • 2019
  • The objective of this study was to evaluate the effects of L-arginine supplementation on muscle damage and fatigue indices and athletic performance improvement of canoe athletes after conducting a high-intensity training program. To achieve the objective, this study applied a high-intensity training program to seven high school canoe athletes. The high-intensity training program is composed of aerobic exercise sessions (twice per week; Tuesday and Thursday), anaerobic exercise sessions (three times per week; Monday, Wednesday, and Friday), and flexibility exercise sessions (five times per week). During the 6 week high-intensity training program, drug ingestion (L-arginine or placebo) was conducted in the first two weeks, wash out (two weeks) followed it, and drug ingestion (L-arginine or placebo) was carried out again in the last two weeks. The crossover design was used for the experiment so all study subjects were assigned to either the L-arginine intake group (the treatment group) or the placebo group (the control group). Each subject ingested 3g per day. This study confirmed the significant effects of L-arginine supplementation on muscle damage indices, fatigue indices, and antioxidants using blood samples. Additionally, FMD was analyzed to evaluate vascular endothelial cell functions and canoe performance was examined using the canoe ergometer. The results of this study showed that L-arginine intake did not have direct effects on the levels of ammonia, IP, and CK. The level of LDH decreased significantly more in the ARG group than in the PLA group due to L-arginine supplementation. Moreover, L-arginine supplementation did not change total NO, d-ROMs, BAP, and FMD significantly. Lastly, the results of the 500m canoe ergometer, which was conducted to evaluate the canoe performance, revealed that L-arginine did not have direct effects on total time, stroke distance, and mean velocity. However, L-arginine supplementation significantly improved muscle damage indices, fatigue indices, antioxidants, FMD, and canoe performance. Therefore, it is believed that additional studies are needed for examining the potential effects of L-arginine supplementation athletic performance enhancement.

Efficient Transformation Method of Soybean Using Meristematic Tissues of Germinating Seeds (발아종자의 분열조직을 이용한 효율적인 콩 형질전환 방법)

  • Kim, Yul-Ho;Park, Hyang-Mi;Choi, Man-Soo;Sohn, Soo-In;Shin, Dong-Bum;Lee, Jang-Yong
    • Korean Journal of Breeding Science
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    • v.40 no.3
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    • pp.278-285
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    • 2008
  • An efficient transformation method for soybean [Glycine max (L.) Merr.] using meristematic tissues of germinating seeds has been established. The embryonic axes were excised from germinating seeds of Korean soybean cultivar, Iksannamulkong and 0.5-2 cm long segment containing meristematic tissues were prepared by cutting hypocotyl region. The explants were inoculated with Agrobacterium tumefaciens strain LBA4404 harboring a binary vector with the bar gene as a selectable marker gene and a ${\beta}-glucuronidase$ (GUSINT) reporter gene, and then co-cultured for 7 days on co-cultivation medium (CCM). The meristematic tissues were cultured on shoot induction medium (SIMP6) supplemented with 0.4 mg/l $N_6-benzylaminopurine$ (BAP) and 0.1 mg/l indolebutyric acid (IBA) in the presence of 6 mg/l L-phosphinotricin (PPT) for 2 weeks and the surviving explants were transferred to shoot elongation medium (SEMP6). Transformation was confirmed by Southern blot analysis and the transformation efficiencies ranged from 1.48 to 2.07%. The new modified transformation method was successfully implemented for obtaining several transgenic lines with SMV-CP gene. It is expected that this method could efficiently be used for the transformation of recalcitrant soybean cultivars.