• 제목/요약/키워드: microplate-based screening

검색결과 5건 처리시간 0.02초

막과산화를 신속히 유발하는 제초제의 고효율 대량스크리닝을 위한 형광검정법 (Fluorescence Assay for High Efficient Mass Screening of the Herbicides Inducing Rapid Membrane Peroxidation)

  • 김진석;권옥경
    • Weed & Turfgrass Science
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    • 제4권4호
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    • pp.308-314
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    • 2015
  • 본 연구는 식물세포막을 파괴시켜 제초활성을 나타내게 하는 화합물(막과산화형 제초제)을 미지의 많은 화합물로부터 신속하게 탐색하기 위한 새로운 검정법을 확립하기 위하여 실시되었으며, 확립한 전체적인 검정과정은 다음과 같다. 96-well microplate에 시험용액 $200{\mu}L$ 넣고 여기에 오이자엽으로부터 적출한 직경 4 mm의 절편 1개씩을 띄운다. 항온실의 광조건 하에서 회전진탕기로 조금씩 흔들어주면서 8시간 배양한 후 절편을 제거한 다음, 배양액에 HVA와 HRP를 첨가하여 반응시킨 후 마이크로평판용 형광검출기를 이용하여 형광도(Ex 320 nm, Em 425 nm)를 측정한다. 형광변화량이 높을수록 제초활성이 높은 것으로 판단한다. 본 방법은 96-well microplate에서 작업을 수행할 수 있고 형광검출 기술을 이용함으로써 검정과정과 작업을 간편하게 하여 검정효율을 기존보다 현저히 높인 것이 특징이다. 아울러 활성검정을 추출된 효소가 아닌 잎 절편수준에서 수행하기 때문에 보다 실용화에 근접한 정량적 데이터를 얻을 수 있는 장점을 가진다.

Towards a Miniaturized Culture Screening for Cellulolytic Fungi and Their Agricultural Lignocellulosic Degradation

  • Arnthong, Jantima;Siamphan, Chatuphon;Chuaseeharonnachai, Charuwan;Boonyuen, Nattawut;Suwannarangsee, Surisa
    • Journal of Microbiology and Biotechnology
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    • 제30권11호
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    • pp.1670-1679
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    • 2020
  • The substantial use of fungal enzymes to degrade lignocellulosic plant biomass has widely been attributed to the extensive requirement of powerful enzyme-producing fungal strains. In this study, a two-step screening procedure for finding cellulolytic fungi, involving a miniaturized culture method with shake-flask fermentation, was proposed and demonstrated. We isolated 297 fungal strains from several cellulose-containing samples found in two different locations in Thailand. By using this screening strategy, we then selected 9 fungal strains based on their potential for cellulase production. Through sequence-based identification of these fungal isolates, 4 species in 4 genera were identified: Aspergillus terreus (3 strains: AG466, AG438 and AG499), Penicillium oxalicum (4 strains: AG452, AG496, AG498 and AG559), Talaromyces siamensis (1 strain: AG548) and Trichoderma afroharzianum (1 strain: AG500). After examining their lignocellulose degradation capacity, our data showed that P. oxalicum AG452 exhibited the highest glucose yield after saccharification of pretreated sugarcane trash, cassava pulp and coffee silverskin. In addition, Ta. siamensis AG548 produced the highest glucose yield after hydrolysis of pretreated sugarcane bagasse. Our study demonstrated that the proposed two-step screening strategy can be further applied for discovering potential cellulolytic fungi isolated from various environmental samples. Meanwhile, the fungal strains isolated in this study will prove useful in the bioconversion of agricultural lignocellulosic residues into valuable biotechnological products.

Estimating dehalogenation reactivity of nanoscale zero-valent iron by simple colorimetric assay by way of 4-chlorophenol reduction

  • Mines, Paul D.;Kaarsholm, Kamilla M.S.;Droumpali, Ariadni;Andersen, Henrik R.;Hwang, Yuhoon
    • Environmental Engineering Research
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    • 제25권2호
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    • pp.197-204
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    • 2020
  • A number of different nanoscale zero-valent iron (nZVI) materials have been prepared and compared depending on the desired properties for the particular application, but different physicochemical properties of this prepared nZVI make it difficult to universally compare and standardize them to the same scale. In this study, we aimed to demonstrate a simple microplate-based colorimetric assay using 4-chlorophenol as an indicator with respect to the remediation of real treatment targets, such as trichloroethylene (TCE), 1,1,1-trichloroethane (TCA), and atrazine. Effect of nickel contents on 4-chlorophenol reduction was successfully investigated by the miniaturized colorimetric assay. In the same manner, the effect of nickel contents on dehalogenation of TCE, TCA, and atrazine was investigated and the pseudo-first-order kinetic constants were compared with the results for 4-chlorophenol. The similar pattern could be observed between 4-chlorophenol reduction obtained by colorimetric assay and TCE, TCA, atrazine reduction obtained by a traditional chromatographic method. The reaction kinetics does not match perfectly, but the degree of reaction can be estimated. Therefore, the colorimetric assay can be a useful and simple screening tool to determine nZVI reactivity toward halogenated organics before it is applied to a particular remediation site.

An International Collaborative Program To Discover New Drugs from Tropical Biodiversity of Vietnam and Laos

  • Soejarto, Djaja D.;Pezzuto, John M.;Fong, Harry H.S.;Tan, Ghee Teng;Zhang, Hong Jie;Tamez, Pamela;Aydogmus, Zeynep;Chien, Nguyen Quyet;Franzblau, Scott G.;Gyllenhaal, Charlotte;Regalado, Jacinto C.;Hung, Nguyen Van;Hoang, Vu Dinh;Hiep, Nguyen Tien;Xuan, Le Thi;Hai, Nong Van;Cuong, Nguyen Manh;Bich, Truong Quang;Loc, Phan Ke;Vu, Bui Minh;Southavong, Boun Hoong
    • Natural Product Sciences
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    • 제8권1호
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    • pp.1-15
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    • 2002
  • An International Cooperative Biodiversity Group (ICBG) program based at the University of Illinois at Chicago initiated its activities in 1998, with the following specific objectives: (a) inventory and conservation of of plants of Cuc Phuong National Park in Vietnam and of medicinal plants of Laos; (b) drug discovery (and development) based on plants of Vietnam and Laos; and (c) economic development of communities participating in the ICBG project both in Vietnam and Laos. Member-institutions and an industrial partner of this ICBG are bound by a Memorandum of Agreement that recognizes property and intellectual property rights, prior informed consent for access to genetic resources and to indigenous knowledge, the sharing of benefits that may arise from the drug discovery effort, and the provision of short-term and long-term benefits to host country institutions and communities. The drug discovery effort is targeted to the search for agents for therapies against malaria (antimalarial assay of plant extracts, using Plasmodium falciparum clones), AIDS (anti-HIV-l activity using HOG.R5 reporter cell line (through transactivation of the green fluorescent protein/GFP gene), cancer (screening of plant extracts in 6 human tumor cell lines - KB, Col-2, LU-l, LNCaP, HUVEC, hTert-RPEl), tuberculosis (screening of extracts in the microplate Alamar Blue assay against Mycobacterium tuberculosis $H_{37}Ra\;and\;H_{37}Rv),$ all performed at UIC, and CNS-related diseases (with special focus on Alzheimer's disease, pain and rheumatoid arthritis, and asthma), peformed at Glaxo Smith Kline (UK). Source plants were selected based on two approaches: biodiversity-based (plants of Cuc Phuong National Park) and ethnobotany-based (medicinal plants of Cuc Phuong National Park in Vietnam and medicinal plants of Laos). At mc, as of July, 2001, active leads had been identified in the anti-HIV, anticancer, antimalarial, and anti- TB assay, after the screening of more than 800 extracts. At least 25 biologically active compounds have been isolated, 13 of which are new with anti-HIV activity, and 3 also new with antimalarial activity. At GSK of 21 plant samples with a history of use to treat CNS-related diseases tested to date, a number showed activity against one or more of the CNS assay targets used, but no new compounds have been isolated. The results of the drug discovery effort to date indicate that tropical plant diversity of Vietnam and Laos unquestionably harbors biologically active chemical entities, which, through further research, may eventually yield candidates for drug development. Although the substantial monetary benefit of the drug discovery process (royalties) is a long way off, the UIC ICBG program provides direct and real-term benefits to host country institutions and communities.

Immunoblot 법을 이용한 간흡충항원(肝吸蟲抗原)의 발육단계별(發育段階別) 항원성분석(抗原性分析)에 관한 연구(硏究) (Studies on the Immunoblot Characterization of Clonorchis sinensis Worm Antigens at Carly Development Stages)

  • 이선경;주경환;정명숙;임한종
    • 농촌의학ㆍ지역보건
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    • 제16권1호
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    • pp.61-69
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    • 1991
  • Serodiagnosis of Clonorchis sinensis infections will probably be a first choice tool for screening of clonorchiasis in a future because of increasing difficulties in collection and examination of stools. The sensitive test such as ELISA can he used effectively. However there are some limitations in serological diagnosis for the detection of serum antibody. One of the major problems is the non-specificity of the antigens which produce cross reaction with other helminthic infection sera. To solve this problem. many investigators have tried to purify the antigens used. In this study, we determined the antigenic profile of the crude saline extract antigen of C. sinensis at early developmental stage based on SDS-PAGE and immunoblotting techniques for the purpose of understanding the nature of C. sinensis worm antigen The following results were obtained : 1) The SDS-PAGE showed many protein hands ranging from 10Kd to 91Kd relative molecular weight. Among them, 66, 46, 40, 33, 27, 24, 16, 14 and 10Kd bands were observed as a principle bands. The protein components of C. sinensis changed chronologically during their early developmental period. 44Kd band was stained unclearly in antigen of 2 weeks worm, but changed to concentrated state in antigen of 5 weeks worm. 35Kd band was found in antigen of 2 weeks worm, however this band was disappeared in antigen of 5 weeks worm. 22Kd band also lost its staining property gradually. 2) In spite of differences in antigenic profile, there was no differences in the data obtained by microplate ELISA using each antigen preparation. Absorbance value began to rise in between 2 to 3 weeks after infection. 3) By EITB. serum antibody recognized major protein bands with molecular weight of 91, 85, 63, 46, 40, 33, 24, 14 and 10Kd hand respectively. Among them 66, 33, 17 and 14Kd bands were observed as non-specific band because they reacted even in normal control sera. Generally, gradual increase of positive reactions were observed as the infection period of C. sinensis was prolonged. In other hand, the reaction of 10Kd hand did not occurred when 26th week sera was tested. 4) The positive reactions using antigens of 2 weeks worm, especially on 40 and 24Kd bands, were most strong and sharply demarcated compared to those of 3~5 weeks worm antigen.

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