• Title/Summary/Keyword: In vitro screening

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Screening of Seed Treatment Fungicide for Control of Damping-off caused by Rhizoctonia solani on Panax ginseng (Rhizoctonia solani에 의한 인삼 모잘록병 방제용 종자처리 약제 선발)

  • Shin, Jeong-Seop;Lee, Seung-Hwan;Cho, Hye-Sun;Cho, Dae-Hui;Kim, Kyoung-Ju;Hong, Tae-Kyun;Park, Chol-Soo;Lee, Seong-Kye;Jung, Hee-Young
    • The Korean Journal of Pesticide Science
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    • v.19 no.4
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    • pp.424-427
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    • 2015
  • Damping-off is a critical disease on ginseng seedling, which caused by the fungal pathogen Rhizoctonia solani. The disease has been prevented by tolclofos-methyl for the last 20 years. However, the tolclofos-methyl usually detected on the harvested roots of 6-year-old ginseng. Herein, we tried to select an alternative pesticide which not only must be safe but also efficiently inhibits the fungal pathogen. Four fungicides (fludioxonil, flutolanil, pencycuron, and thifluzamide) were applied to their inhibition efficacy against the pathogen. In in vitro test, fludioxonil treatment showed 80% inhibition activity for 25 days. Thifluzamide and flutolanil showed the activity for 10 days. Pencycuron showed for 1 days. In addition, the fludioxonil was more effective to control the pathogen comparing to other three fungicides in field. The incidence of damping-off was reduced to 71% by fludioxonil treatment. The level of the fungicide residue in seedling was 0.44 mg/kg, which value will be a negligible level in final products after 5 years. Consequently, the fludioxonil is a conceivable alternative for tolclofos-methyl to cope with R. solani.

Identification of the Oligotrophic Bacteria Strain 7F Biocontrolling Phytophthora Blight Disease of Red-pepper (고추 역병 방제를 위한 저영양 길항세균 7F 균주의 동정)

  • Kim, Dong-Gwan;Yeo, Yun-Soo;Kwon, Soon-Wo;Jang, Kil-Su;Lee, Chang-Muk;Lee, Mi-Hye;Kim, Soo-Jin;Koo, Bon-Sung;Yoon, Sang-Hong
    • Research in Plant Disease
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    • v.16 no.1
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    • pp.41-47
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    • 2010
  • A total of 10,753 oligotrophic bacteria were isolated from the cultivated soils of red-pepper infected by Phytophthora blight disease in various regions of Korea (Chungju, Anmyon, Taean, Andong, Eumsung and Goesan). Seven bacteria isolates among these collected resources were selected by the first screening of in vitro antagonistic assay against major several plant pathogenic fungi including Phytophthora capsici. Finally, strain 7F was selected by pot assay for a possible biological control agent against Phytophthora blight disease of pepper seedling in the greenhouse. Strain 7F was identified as Bacillus subtilis on the basis of its 16S rDNA sequence analysis and as standardized biochemical characteristics assay kits such as API20 NE. In the experiment of P. capsici zoospore infected red-pepper on the pot test, infection rate of red-pepper with nonetreatment to Phytophthora blight disease was 87%, while the rate was only 6% in the pot treated with strain 7F. This result indicated that the Bacillus subtilis strain 7F will be useful as a potential biocontrol agent for Phytophthora blight disease of red-pepper.

A Study on the Antioxidative, Anti-inflammatory and Anti-thrombogenic Effects of Zanthoxylum piperitum DC. Extract (초피나무 추출물의 항산화, 항염증 및 항혈전 효능에 관한 연구)

  • Jang, Mi-Jin;Rhee, Soon-Jae;Cho, Sung-Hee;Woo, Mi-Hee;Choi, Jeong-Hwa
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.35 no.1
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    • pp.21-27
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    • 2006
  • Effects of root, stem and leaf extracts of Zanthoxylum piperitum on the inhibition of lipid peroxidation in the hepatic microsome of rat, DPPH radical scavenging activity, soybean lipoxygenase activity and activated partial thromboplastin times (APTT) were examined in vitro. The highest inhibition of hepatic microsomal lipid peroxidation was observed by ethyl acetate fraction of the root and stem extracts. The high inhibition of lipid peroxidation was observed in the leaf, the root and the stem in order. The DPPH radical scavenging activity of ethyl acetate fraction was higher than that of n-butanol fraction and it was similar to the root and the steam extract. It was similar to the inhibition of hepatic microsomal lipid peroxidation. The DPPH radical scavenging activity was the highest in 0.50 mg/mL of ethyl acetate fraction, and it was 4.4-fold higher than that of a-tocopherol, as an antioxidant standard. The DPPH radical scavenging activity was dependent on the extract concentration in the range of $0.12\~5.00$ mg/mL. The soybean lipoxygenase activity of ethyl acetate fraction was higher than that of n-butanol fraction and it was similar to the root and the stem extracts. The soybean lipoxygenase activity was the highest in 0.50 mg/mL of ethyl acetate fraction. The soybean lipoxygenase activity was dependent on the extract concentration in the range of $0.12\~5.00$ mg/mL. The leaf extract showed the highest antithrombogenic effect followed by the stem and then the root extract. The activated partial thromboplastin times were dependent on the extract concentration in the range of $0.10\~2.00$ mg/mL.

A novel anti-adhesion agent using DNA aptamers for Streptococcus mutans (DNA 앱타머를 이용한 Streptococcus mutans의 부착 억제)

  • Park, Byung-Ju;Ohk, Seung-Ho
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.19 no.3
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    • pp.382-388
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    • 2018
  • In this study, the SELEX method was used to screen for and select aptamers with high selectivity and affinity for Streptococcus mutans, which is the causative agent of dental caries. Aptamers are single stranded oligonucleotides of DNA or RNA with high selectivity and affinity for target molecules because of their specific three-dimensional structures that are used to collect biometric information. When compared to antibodies in vitro, aptamers are more advantageous because of their ease of use in the screening process, low cost, chemical stability, and lack of restrictions on the target molecules. We sorted DNA aptamers, which contain 44 bp or 38 bp primer sequences in 5' and 3', 39 bp random sequences in the middle.We then analyzed the character and affinity to S. mutans. Aptamers of specific oligonucleotides that combine with S. mutans were selected and these results are selectively fused to S. mutans. The results confirmed that DNA aptamers can be used for rapid diagnosis and treatment of dental caries caused by bacteria of the oral cavity, namely S. mutans.

Selection of a Triploid Poplar by Flow Cytometric Analysis and Growth Characteristics of its in vitro Grown Plants (유세포 분석을 통한 현사시나무 3배체 선발 및 계통별 기내생장 특성)

  • Bae, Eun-Kyung;Lee, Hyoshin;Lee, Jae-Soon;Noh, Eun-Woon
    • Journal of Korean Society of Forest Science
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    • v.101 no.2
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    • pp.291-296
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    • 2012
  • Triploids are a useful tool for biomass production and molecular breeding of trees with a long life span. Triploids of the poplar 'Hyunsasi' (Populus alba ${\times}$ P. glandulosa) have been developed by crossing between female diploids and a male tetraploid. The tetraploid was developed around the 1970s at Korea Forest Research Institute by colchicine-induced chromosome doubling. Seedlings of the $F_1$ generation were analyzed using flow cytometry to verify their ploidy status. The mean relative fluorescence index of 3 F1 poplars, labeled as Line- 1, Line-17, Line-18, were approximately 1.5 times higher than those of diploid poplars, and the results clearly indicated that they were triploids. The phenotype of the F1 poplars included larger leaves and thicker stem than diploids, and abnormal leaf morphology, especially in the triploid 'Line-18'. Three triploid lines developed roots more slowly and had less roots than diploid. However, 3 poplar cytotypes (2x, Line-1, Line-17) rooted within 10 days on MS medium. In contrast, compared with the 3 cytotypes, the Line-18 showed about 80% and 70% in the rooting rate and the number of roots. The triploid poplars could be directly utilized for biomass production and with their sterility, they could serve as basic material for genetic transformation. In addition, flow cytometric analysis proved to be an effective and reliable method for screening forest trees for their ploidy level.

Culture Conditions of E. coli Harboring Human O-Linked N-Acetyl-${\beta}$-Glucosaminidase Gene and Enzymatic Properties (사람의 O-linked-N-acetyl-${\beta}$-D-glucosaminidase 유전자를 함유한 대장균의 배양조건과 효소학적 특성)

  • 강대욱;조용권;서현효
    • Korean Journal of Microbiology
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    • v.40 no.2
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    • pp.147-153
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    • 2004
  • Protein modification by N-acetyl-${\beta}$-D-glucosamine (O-G1cNAc) on the hydroxyl groups of Ser or Thr ubiq-uitously occurs in eukaryotic cells and is involved in many cellular phenomena. The level of O-G1cNAc-mod-ified protein is regulated by OGT and O-GlcNAcase enzymes. We have tried to produce recombinant O-GlcNAcase in E. coli as an effort to establish in vitro screening system for modulators of O-GlcNAcase. The culture conditions for improvement of O-GlcNAcase productivity, were as follows: induction temperature, $30^{\circ}C$; the concentration of L-arabinose, 0.02% and induction time, 5 hr. Under these culture conditions, E. coli cells containing O-GlcNAcase gene had no enzyme activity until up to 3 hr culture. However, O-GlcNAcase activity dramatically increased from 3 to 5 hr culture. It almost maintained the same level after 5 hr culture. Western blot analysis verified the amount of expressed O-GlcNAcase increased with culture time, being con-sistent with activity data. The optimal reaction condition determined in this study was as follows: protein quan-tity, $5{\mu}g$; reaction time, 30 min; reaction temperature, $45^{\circ}C$; substrate concentration, 2 mM; reaction pH, 6.5. Methanol had little effect on O-GlcNAcase activity and 90% of activity were retained at 10%. Only 15% resid-ual activity were detected at 5% of chloroform.

Screening for Effective Organic Farming Materials for the Control of Cucumber Scab Caused by Cladosporium cucumerinum (오이 검은별무늬병 방제에 효과적인 유기농업자재 선발)

  • Park, So-Hyang;Hong, Sung-Jun;Shim, Chang-Ki;Kim, Min-Jeong;Park, Jong-Ho;Han, Eun-Jung;Kim, Yong-Ki
    • Research in Plant Disease
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    • v.23 no.2
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    • pp.159-167
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    • 2017
  • This study was conducted to evaluate the control efficacy of the organic farming materials on cucumber scab caused by Cladosporium cucumerinum PT1 (KACC 48094). The antifungal activities in vitro as well as the suppressive effect of 43 organic farming materials on the spore germination and germ tube growth by inoculating spore suspension on cucumber seedlings in vivo were investigated. Thirteen organic farming materials inhibited the mycelial growth of C. cucumerinum and nine of these were microbial agents. In the screening using cucumber seedlings, six organic farming materials were very effective with control efficacy value of 90%. Among them, Bacillus amyloliquefaciens M27 provided suppressive effect on both mycelial growth and spore germination against cucumber scab. Finally, nine organic farming materials were selected to test the protective and curative effects, and all chosen organic farming materials significantly suppressed disease incidence when applied in the preventive action, in comparison with the curative action. Especially, Bordeaux mixture I and III gave excellent protective control efficacy with control values of 96.7% and 73.3%, respectively, whereas its curative control effect was significant low. Among these, only Thymus quinquecostatus+Sophora extract showed curative activity, although the control value was as low as 50%. This study suggests that cucumber scab can be controlled by some organic farming materials in the farmhouses under comparatively cold and wet condition and protective treatment is more important and efficient.

Screening of Flavonoid Compounds with HMG-CoA Reductase Inhibitory Activities (플라보노이드 화합물로부터 HMG-CoA reductase 저해 활성 물질 탐색)

  • Son, Kun Ho;Lee, Ju Yeon;Lee, Jeong Soon;Kang, Sam Sik;Sohn, Ho Yong;Kwon, Chong Suk
    • Journal of Life Science
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    • v.28 no.2
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    • pp.247-256
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    • 2018
  • 3-Hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors (statins) are widely used drugs for lowering blood lipid levels and preventing cardiovascular diseases. HMG-CoA reductase is a key enzyme to control the biosynthesis of cholesterol. We have tested HMG-CoA reductase-inhibitory activity on the flavonoids of 98 species in vitro. The anti-hypercholesterolemic activities of flavonoids were studied using an HMG-CoA reductase assay equipped with a 96-well UV plate. This assay was based on the spectrophotometric measurement of the decrease in absorbance, which represents the oxidation of NADPH by the catalytic subunit of HMG-CoA reductase in the presence of the substrate HMG-CoA. Among the clinically available statins, pravastatin was used as a positive control. Among the tested compounds, kuraridin, morin and sophoraflavanone G showed strong inhibition activities. In particular, morin and sophoraflavanone G inhibited HMG-CoA reductase by 45.0% and 54.6% at a concentration of $10{\mu}g/ml$, and the $IC_{50}$ values were calculated to $13.31{\mu}g/ml$ and $7.26{\mu}g/ml$ respectively.

Characterization of Bacillus amyloliquefaciens DA12 Showing Potent Antifungal Activity against Mycotoxigenic Fusarium Species

  • Lee, Theresa;Park, Dami;Kim, Kihyun;Lim, Seong Mi;Yu, Nan Hee;Kim, Sosoo;Kim, Hwang-Yong;Jung, Kyu Seok;Jang, Ja Yeong;Park, Jong-Chul;Ham, Hyeonheui;Lee, Soohyung;Hong, Sung Kee;Kim, Jin-Cheol
    • The Plant Pathology Journal
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    • v.33 no.5
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    • pp.499-507
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    • 2017
  • In an attempt to develop a biological control agent against mycotoxigenic Fusarium species, we isolated Bacillus amyloliquefaciens strain DA12 from soil and explored its antimicrobial activities. DA12 was active against the growth of mycotoxigenic F. asiaticum, F. graminearum, F. proliferatum, and F. verticillioides both in vitro and in planta (maize). Further screening using dual culture extended the activity range of strain DA12 against other fungal pathogens including Botrytis cinerea, Colletotrichum coccodes, Endothia parasitica, Fusarium oxysporum, Raffaelea quercus-mongolicae, and Rhizoctonia solani. The butanol extract of the culture filtrate of B. amyloliquefaciens DA12 highly inhibited the germination of F. graminearum macroconidia with inhibition rate 83% at a concentration of $31.3{\mu}g/ml$ and 100% at a concentration of $250{\mu}g/ml$. The antifungal metabolite from the butanol extract was identified as iturin A by thin layer chromatography-bioautography. In addition, volatile organic compounds produced by DA12 were able to inhibit mycelial growth of various phytopathogenic fungi. The volatile compounds were identified as 2-heptanone, 5-methyl heptanone and 6-methyl heptanone by gas chromatography-mass spectrometry (GC-MS) analysis. These results indicate that the antagonistic activity of Bacillus amyloliquefaciens DA12 was attributable to iturin A and volatile heptanones, and the strain could be used as a biocontrol agent to reduce the development of Fusarium diseases and mycotoxin contamination of crops.

Effects of Caffeic Acid Methyl Ester from Mentha arvensis Linne var. piperascens on Function of E6 Oncoprotein of HPV 16 (박하에서 분리한 Caffeic Acid Methyl Ester가 자궁경부암 바이러스 발암단백질 E6의 기능에 미치는 영향)

  • Baek, Tae-Woong;Lee , Kyung-Ae;Ahn , Min-Jung;Joo, Hae-Hyung;Oh , Hyun-Cheol;Ahn , Jong-Seok;Cho , Yong-Kweon;Myung , Pyung-Keun;Yoon, Do-Young
    • YAKHAK HOEJI
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    • v.48 no.6
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    • pp.328-334
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    • 2004
  • Cervical cancer is one of the leading causes of female death. Viral oncoproteins E6 and E7 are selectively retained and expressed in carcinoma cells infected with HPV (Human pa pilloma virus) type 16 and cooperated in immotalization and transformation of primary keratinocyte. E6 and E7 oncoproteins interfere the functions of tumor suppressor proteins p53 and retinoblasoma protein (pRb), respectively. Among a lots of natural products, Mentha arvensis Linne var.piperascens have inhibitory effects on bindings between E6 oncoprotein and tumor suppressor p53, E3 ubiqutin- protein ligase (E6AP). HPV oncoprotein inhibitors from Mentha piperita L. were isolated by solvent partition and column chromatography (Silica gel, RP-18) and inhibitory compounds were finally purified by HPLC using an ELISA screening system based on binding between E6 and E6AP. The aim of this study is to identify the structure of inhibitory compounds and to investigate whether these compounds have inhibitory effects on functions of E6 oncoprotein. We investigated whether caffeic acid methyl ester (CAM) extracted from Mentha piperita L. could inhibit the function of E6 oncoprotein. CAM inhibited the in vitro binding of E6 and E6AP which are essential for the binding and degradation of the tumor suppressor p53 and also inhibited the proliferation of human cervical cancer cell lines (SiHa and CaSKi) in a dose response manner. These results suggest that CAM inhibited the function of E6 oncoprotein, suggesting that it can be used as a potential drug for the treatment of cervical cancers infected with HPV.