• Title/Summary/Keyword: integrated T-DNA

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New ecological health assessment approaches of an urban stream using molecular and physiological level biomarkers and bioindicators

  • Kim, Ja-Hyun;Yeom, Dong-Hyuk;Kim, Joon-Ha;An, Kwang-Guk
    • Animal cells and systems
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    • v.16 no.4
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    • pp.329-336
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    • 2012
  • This study evaluated ecological health, using various biomarkers and bioindicators, of pale chub (Zacco platypus) as a sentinel species, in Daejeon Stream, South Korea, during AprilMay 2011. The biomarkers and bioindicators were compared among three sites of control: Reference ($C_z$), transition ($T_z$), and the urban zones ($U_z$); and the 7-Ethoxyresorufin-O-deethylase (EROD) activity, DNA damage, acetylcholinesterase (AChE) activity, and vitellogenin (VTG) concentrations were more significantly increased in the $U_z$ than in the $C_z$. Also, physiological markers such as condition factor, liver somatic index, visceral somatic index, and gonad somatic index were significantly increased in the $U_z$ than in the $C_z$. For the health assessments, three categorized parameters of blood chemistry, molecular biomarkers, and physiological bioindicators were standardized and calculated as a star-plot, representing values of Integrated Health Response (IHR). Values of IHR had more significant (P<0.05) increases in the $U_z$ than any other zones, indicating an impairment of ecological health by organic matter, nutrients (N, P), and toxic chemicals. This study is based on low levels of biological organization approach of molecular and physiological biomarkers and bioindicators, so further study of high-levels of biological organization approach such as community and population is required for overall range of health assessments. The approach of IHR values, however, may be useful in providing early warning of future impacts on ecological health.

Pests occurring on Cymbidium (심비디움에 발생하는 해충의 종류)

  • Cho, Myoung Rae;Jeon, Sung-Wook;Kang, Taek Joon;Kim, Hyung Hwan;Ahn, Seung-Joon;Yang, Chang Yeol
    • Korean journal of applied entomology
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    • v.52 no.4
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    • pp.403-408
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    • 2013
  • A survey of pest occurrence and status of farmer's pest management was conducted at 45 cymbidium farms in 10 major cultivation areas in Korea. The pest species collected from the cymbidium farms were identified as follows: Tetranychus urticae Koch, Frankliniella intonsa Trybom, Pinnaspis aspidistrae Signoret, Incilaria confusa Cockarel, Halyomorpha brevis Walker, Myzus persicae S$\ddot{u}$lzer, and Aphis gossypii Glover, Coccus hesperidum Linnaeus, Thrips flavus Schrank, and Thrips tabaci Lindeman. The two-spotted spider mite, T. urticae, was the key pest in cymbidium production, occurring on 45 farms, followed by scales (20 farms), slugs (6), thrips (8), aphids (5), and stinkbug (1). PCR-RFLP of the rDNA ITS2 region revealed that two thrips species, Thrips flavus Schrank and Thrips tabaci Lindeman, occur on cymbidium farms. Therefore, it is necessary for the cymbidium farmers to establish an integrated pest management system to meet quarantine standards.

Development of High Quality Forage Grass by Down-regulating Lignin Biosynthetic Gene (리그닌 생합성관련 유전자의 발현조절에 의한 고품질 목초 개발)

  • Woo Hyun-Sook;Yun Jung-Woo;Lee Byung-Hyun
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.26 no.1
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    • pp.1-8
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    • 2006
  • To develop a new variety of orchardgrass with improved digestibility, caffeic acid O-methyltransferase (Dgcomt), which is a methylation enzyme involved in the early stages of lignin biosynthesis, was isolated and characterized. Dgcomt was expressed not only in leaves but also in stems and roots. The expression levels of transcripts were high in stems and roots which are the most lignified tissues, and only moderate levels of transcripts were expressed in leaves. To develop transgenic orchardgrass plants by down-regulating the Dgcomt gene, an RNAi suppression vector with partial Dgcomt DNA fragment was constructed and transferred into the genome of orchardgrass via Agrobacterium-mediated gene transfer method. PCR and Southern blot analyses with genomic DNAs from putative transgenic plants revealed that the T-DNA region containing RNAi construct was successfully integrated into the genome of orchardgrass. Northern blot analysis revealed that the majority of the down-regulated transgenic lines showed significant reduction in Dgcomt gene expression. These RNAi transgenic orchardgrass will be useful for molecular breeding of new variety with improved digestibility by down-regulating lignin biosynthetic enzyme.

EST Analysis system for panning gene

  • Hur, Cheol-Goo;Lim, So-Hyung;Goh, Sung-Ho;Shin, Min-Su;Cho, Hwan-Gue
    • Proceedings of the Korean Society for Bioinformatics Conference
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    • 2000.11a
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    • pp.21-22
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    • 2000
  • Expressed sequence tags (EFTs) are the partial segments of cDNA produced from 5 or 3 single-pass sequencing of cDNA clones, error-prone and generated in highly redundant sets. Advancement and expansion of Genomics made biologists to generate huge amount of ESTs from variety of organisms-human, microorganisms as well as plants, and the cumulated number of ESTs is over 5.3 million, As the EST data being accumulate more rapidly, it becomes bigger that the needs of the EST analysis tools for extraction of biological meaning from EST data. Among the several needs of EST analyses, the extraction of protein sequence or functional motifs from ESTs are important for the identification of their function in vivo. To accomplish that purpose the precise and accurate identification of the region where the coding sequences (CDSs) is a crucial problem to solve primarily, and it will be helpful to extract and detect of genuine CD5s and protein motifs from EST collections. Although several public tools are available for EST analysis, there is not any one to accomplish the object. Furthermore, they are not targeted to the plant ESTs but human or microorganism. Thus, to correspond the urgent needs of collaborators deals with plant ESTs and to establish the analysis system to be used as general-purpose public software we constructed the pipelined-EST analysis system by integration of public software components. The software we used are as follows - Phred/Cross-match for the quality control and vector screening, NCBI Blast for the similarity searching, ICATools for the EST clustering, Phrap for EST contig assembly, and BLOCKS/Prosite for protein motif searching. The sample data set used for the construction and verification of this system was 1,386 ESTs from human intrathymic T-cells that verified using UniGene and Nr database of NCBI. The approach for the extraction of CDSs from sample data set was carried out by comparison between sample data and protein sequences/motif database, determining matched protein sequences/motifs that agree with our defined parameters, and extracting the regions that shows similarities. In recent future, in addition to these components, it is supposed to be also integrated into our system and served that the software for the peptide mass spectrometry fingerprint analysis, one of the proteomics fields. This pipelined-EST analysis system will extend our knowledge on the plant ESTs and proteins by identification of unknown-genes.

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NDP-sugar production and glycosylation of ${\varepsilon}$-rhodomycinone in Streptomyces venezuelae (Streptomyces Peucetius에서의 ${\varepsilon}$-rhodomycinone 추출 및 이종균주에서의 rhodomycin D 생산 연구)

  • Park, Sung-Hee;Cha, Min-Ho;Kim, Eun-Jung;Yoon, Yeo-Joon;Sohng, Jae-Kyung;Lee, Hee-Chan;Liou, Kwang-Kyoung;Kim, Byung-Gee
    • KSBB Journal
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    • v.23 no.1
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    • pp.44-47
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    • 2008
  • Anthracycline antibiotics doxorubicin (DXR) is clinically important cancer therapeutic agent produced by Streptomyces peucetius. DXR result by further metabolism of rhodomycin D (RHOD) and require a deoxy-sugar component for their biological activity. In this study, production of TDP-L-daunosamine and its attachment to ${\varepsilon}$-rhodomycinone (RHO) to generate RHOD has been achieved by bioconversion in Streptomyces venezuelae that bears eleven genes. S. peucetius seven genes (dnmUTJVZQS) were transformed by plasmid and S. venezuelae two genes desIII, IV and two more S. peucetius drrA, B genes were integrated into chromosomal DNA. To generate the feeding substrate RHO, 6L S. peucetius grown on agar plate was harvested, extracted with organic solvent and then purified using preparative HPLC. Recombinant S. venezuelae grown on agar plate containing RHO was harvested and its n-butanol soluble components were extracted. The glycosylated product of aromatic polyketide RHO using heterologous host S. venezuelae presents the minimal information for TDP-L-daunosamine biosynthesis and its attachment onto aglycone. Moreover, the structure of auxiliary protein, DnrQ, was predicted by fold recognition and homology modeling in this study. This is a general approach to further expand of new glycosides of antitumor anthracycline antibiotics.

유전공학기법으로 변형시킨 내성유전자네 대한 수질환경에서의 전이동태

  • 이성기;김치경
    • Korean Journal of Microbiology
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    • v.30 no.4
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    • pp.322-331
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    • 1992
  • In order to understand the transfer and behavior of R gene in water environments. the Kmr gene in the genetically modified microorganisms(GMMs) w,is studied by conjugation. The plasmid variously rearranged in the conjugants were comparatively analyzied by agarosc gel electrophoresis and the specific Km' genes in the gel were tletected with DNA probe. The Kmr genes of the GMM strains(DKC600 and DKC601) were transferred at higher rate than those of natural isola~e(DKI)b, ut the ratc was a little diflurent depending upon the recipient strains. Rearrangement of the plasmids appeared morc drastic in GMM strains than in IIKI as donor. The transfer frequencies of the Km' genes in LR broth were remarkably higher than in the water of AW and FW without regards to the strains. In LA breth. the frequencies of Kmr genes were higher at 25'C-30$^{\circ}$C than at 10$^{\circ}$C and at pH - 7 than pH 9, but temperature and pH of the FW did n,,t affect to the frequency. And the conjugants from GMM strains in FW did not showed any plasmids. except tor 43 kb plasmiil. As results of Southern analysis of the plasmid, variously rearranged in eonjugant cells obtained in LB broth, the Kmr genes were detected at the same position of Km' plasrnids of the donor cell(DK1 and GMM strains). But Km' plasmid disappeared in the conjugants obtained in F'W and their chronlosomes showed strong signal of hybridization. The Kmr plasmid of DKl in the conjugants obtained in FW water was transferred and maintained its size, but the Kmr plasinids of the GMM strains were all integrated into chromosome. Therefore, the Kmr plasmids of DKI anit GMM strains in LH were intactly transferred and other plasmitls were variously rearranged. but Km' gene of DKC600 in FW water was integrated into the chromosorn: without regards to the temperature and pH of the water.

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Effect of Callus Type and Antioxidants on Plant Regeneration and Transformation of Tall Fescue (캘러스의 형태와 항산화물질 첨가가 톨 페스큐의 식물체 재분화와 형질전환효율에 미치는 영향)

  • Lee Ki-Won;Lee Sang-Hoon;Kim Do-Hyun;Lee Dong-Gi;Won Sung-Hye;Lee Hyo-Shin;Lee Byung-Hyun
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.26 no.2
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    • pp.77-82
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    • 2006
  • An efficient transformation system for the production of transgenic plants has been developed for tall fescue (Festuca arundinacea Schreb.) via Agrobacterium-mediated transformation of seed-derived callus. From the point of morphogenetic capacity, three types of callus were selected. High frequency of plant regeneration was obtained by selection of type II callus, and the plant regeneration frequency was 52.6% when embryogenic callus were cultured on the regeneration medium. Supplementation of the media with 10 mg/L $AgNO_3$ and 40 mg/L cysteine enhanced frequencies of plant regeneration up to 65.3%. The highest transformation efficiency was also obtained when type II callus were inoculated with Agrobacterium. Southern blot analysis of PCR products of transgenic plants demonstrated that transgenes were successfully integrated into the genome of tall fescue. Efficient regeneration system and transformation established in this study will be useful for molecular breeding of tall fescue through genetic transformation.

Inhibition of in Vitro Growth of Three Soil-borne Turfgrass Diseases by Antagonistic Bacteria from Composted Liquid Manure (가축분뇨액비의 길항미생물에 의한 토양전염성 병원균의 생육억제 효과)

  • Ryu, Ju Hyun;Shim, Gyu Yul;Kim, Ki Sun
    • Horticultural Science & Technology
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    • v.32 no.6
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    • pp.879-886
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    • 2014
  • This study was conducted to test in vitro the antagonistic effect of composted liquid manure (CLM) against soil-borne turfgrass pathogenic fungi, Rhizoctonia solani AG-2-2 (IIIB) (brown patch), R. solani AG-2-2 (IV) (large patch), and Sclerotinia homoeocarpa (dollar spot) for environmentally friendly turfgrass management. CLMs were collected from 9 livestock excretion treatment facilities around the country including Gunwi (GW), Hapcheon (HC), Hoengseong (HS), Icheon (IC), Iksan (IS), Muan (MA), Nonsan (NS), and Yeoju (YJ). CLMs of IC, GW, and IS showed s ignificant (p < 0.05) mycelium growth inhibition that was 17.8%, 20.4%, and 48.0% against R. solani AG-2-2 (IIIB), R. solani AG-2-2 (IV), and S. homoeocarpa, respectively. A t otal of 110 bacterial isolates were obtained from the CLMs that showed antagonistic effects. Among them, 5, 4, and 10 microbe isolates showed promising antifungal activity against mycelium growth of R. solani AG-2-2 (IIIB), R. solani AG-2-2 (IV), and S. homoeocarpa, respectively. The bacterial isolates ICIIIB60, GWIV70, and ISSH20 effectively inhibited the mycelial growth of three soil-borne turfgrass pathogens. Selected bacterial isolates were identified as Alcaligenes sp., Bacillus licheniformis Ab2, and B. subtilis C7-3 through 16s rDNA gene sequence analysis. Among 5 fungicides, the most compatible fungicide with ICIIIB60, GWIV70, and ISSH20 was tebuconazol, toclofos-methyl and toclofos-methyl, respectively. These findings suggested that CLMs could be effectively used not only as organic liquid fertilizer sources but also as biological control agents for soil-borne turfgrass diseases such as brown patch, large patch, and dollar spot.

Cytological Study of the Introduction of Agrobacterium tumefaciens Spheroplasts into Nicotiana tabacum Protoplasts (Agrobacterium tumefaciens Spheroplast의 연초엽육 Protoplast내 도입에 관한 세포학적 연구)

  • Kim, Jung-Hye;Koo, Yong-Bum;Lee, Ki-Yung
    • Journal of Yeungnam Medical Science
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    • v.2 no.1
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    • pp.175-181
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    • 1985
  • Agrobacterium tumefaciens induces cancerous growths called crown galls at wound sites on dicotyledonous plants. A large plasmid called Ti plasmid is responsible for virulence. Upon tumor induction, part of the plasmid, termed T-DNA, becomes integrated into plant genome and its genetic sequences are expressed. These properties allow Ti plasmids to be used as gene vectors in plants. Several in vitro methods for the transfer of Ti plasmid into plant cell have been developed. One of them is the treatment of bacterial spheroplasts and plant protoplasts mixture with polyethylene glycol that is generally used as fusogen in cell-to-cell fusion. Several workers investigated the interaction of bacterial spheroplasts with plant protoplasts in the presence of polyethylene glycol and suggested that the interaction is not fusion but endocytosis. In this report we observed the interaction of Agrobacterium tumefaciens spheroplasts with Nicotiana tabacum protoplasts by electron microscope. Agrobacterium tumefaciens spheroplasts and Nicotiana tabacum protoplasts were prepared and mixed in the presence of polyethylene glycol and high pH-high $Ca^{2+}$ buffer. Then the interaction of the spheroplasts with the protoplasts was examined by transmission electron microscope. After the treatment of polyethylene glycol the spheroplasts adhered to the surface of the protoplasts and then they were engulfed by the protoplasts. After the high pH-high $Ca^{2+}$ buffer treatment the engulfed spheroplasts lost their cell integrity. No fusion process was observed. Thus all these observations suggest that the introduction process of Agrobacterium tumefaciens spheroplasts into Nicotiana tabacum protoplasts with the aid of polyethylene glycol is endocytosis.

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Effect of SeaR gene on virginiamycins production in Streptomyces virginiae (희소방선균 SeaR 유전자가 Streptomyces virginiae의 virginiamycins 생산에 미치는 영향)

  • Ryu, Jae-Ki;Kim, Hyun-Kyung;Kim, Byung-Won;Kim, Dong-Chan;Lee, Hyeong-Seon
    • Korean Journal of Microbiology
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    • v.51 no.3
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    • pp.256-262
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    • 2015
  • In order to study the effect of the receptor protein (SeaR), which is isolated from Saccharopolyspora erythraea, we introduced the SeaR gene to Streptomyces virginiae as host strains. An effective transformation procedure for S. virginiae was established based on transconjugation by Escherichia coli ET12567/pUZ8002 with a ${\varphi}C31$-derived integration vector, pSET152, which contained int, oriT, attP, and $ermEp^{\ast}$ (erythromycin promotor). Therefore, the pEV615 was introduced into S. virginiae by conjugation and integrated at the attB locus in the chromosome of the recipients by the ${\varphi}C31$ integrase (int) function. Transformants of S. virginiae containing the SeaR gene were confirmed by PCR and transcriptional expression of the SeaR gene in the transformants was analyzed by RT-PCR, respectively. And, we examined the production time of virginiamycins in the culture media of both the transformants and the wild type. The production time of virginiamycins in the wild type and transformants was the same. When 100 ng/ml of synthetic $VB-C_6$ was added to the state of 6 or 8 hour cultivation of wild type and transformants, respectively, the virginiamycins production was induced, meaning that the virginiamycins production in the wild type was detected 2 h early than transformants. From these results, SeaR expression was also affected to virginiamycins production in transformants derived from S. virginiae. In this study, we showed that the SeaR protein worked as a repressor in transformants.