• Title/Summary/Keyword: Cell suspension

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Bacillus spp. as Biocontrol Agents of Root Rot and Phytophthora Blight on Ginseng

  • Bae, Yeoung-Seuk;Park, Kyungseok;Kim, Choong-Hoe
    • The Plant Pathology Journal
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    • v.20 no.1
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    • pp.63-66
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    • 2004
  • Ginseng (Panax ginseng) is one of the most widely cultivated medicinal herbs in Korea. However, yield losses reached up to 30-60% due to various diseases during 3 or 5 years of ginseng cultivation in the country. Therefore, successful production of ginseng roots depends primarily on the control of diseases. The objective of this study was to select potential biocontrol agents from rhizobacteria isolated from various plant internal root tissues for the control of multiple ginseng diseases as an alternative to fungicides. Among 106 Bacillus strains, two promising biocontrol agents, Bacillus pumilus strain B1141 and Paenibacillus lentimobus strain B1146, were selected by screening against root rot of ginseng caused by Cylindrocarpon destructans in a greenhouse. Pre-inoculation of selected isolates to seed or l-year-old root of ginseng resulted in stimulation of shoot and/or root growth of seedlings, and successfully controlled root rot caused by C. destructans (P<0.05). Furthermore, drenching of cell suspension of the selected isolates on seedling-growing pots reduced the incidence of Phytophthora blight after the seedlings were challenged with zoospores of Phytophthora cactorum (P<0.05). P. lentimorbus strain B1146 showed antifungal activity against various soil-borne pathogens in vitro, while B. pumilus strain B1141 did not show any. Results of this study suggest that some rhizobacteria can induce resistance against various plant diseases on ginseng.

Validation and Application of a Real-time PCR Protocol for the Specific Detection and Quantification of Clavibacter michiganensis subsp. sepedonicus in Potato

  • Cho, Min Seok;Park, Duck Hwan;Namgung, Min;Ahn, Tae-Young;Park, Dong Suk
    • The Plant Pathology Journal
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    • v.31 no.2
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    • pp.123-131
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    • 2015
  • Clavibacter michiganensis subsp. sepedonicus (Cms) multiplies very rapidly, passing through the vascular strands and into the stems and petioles of a diseased potato. Therefore, the rapid and specific detection of this pathogen is highly important for the effective control of the pathogen. Although several PCR assays have been developed for detection, they cannot afford specific detection of Cms. Therefore, in this study, a computational genome analysis was performed to compare the sequenced genomes of the C. michiganensis subspecies and to identify an appropriate gene for the development of a subspecies-specific PCR primer set (Cms89F/R). The specificity of the primer set based on the putative phage-related protein was evaluated using genomic DNA from seven isolates of Cms and 27 other reference strains. The Cms89F/R primer set was more specific and sensitive than the existing assays in detecting Cms in in vitro using Cms cells and its genomic DNA. This assay was also able to detect at least $1.47{\times}10^2copies/{\mu}l$ of cloned-amplified target DNA, 5 fg of DNA using genomic DNA or $10^{-6}$ dilution point of 0.12 at $OD_{600}$ units of cells per reaction using a calibrated cell suspension.

Effect of Herbicide Paraquat on NAD(H)-Redox-cycle (제초제 Paraquat의 NAD(H) 산화환원에 대한 영향)

  • Kim Mi-Lim;Choi Kyung-Ho
    • Journal of Life Science
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    • v.15 no.2 s.69
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    • pp.304-310
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    • 2005
  • This study was carried out to investigate the effect of herbicide paraquat (1,1-dimethyl-4,4-bipyridilium dichloride) on the electron transport system of the cell. When actively growing cells of bacteria were exposed to the 1.0 mM paraquat, more than $50\%$ of the cells were killed at 0 hour. But specific activities of superoxide dismutase (SOD) were not changed at 0 hour of paraquat treatment. Oxido-reductions of NAD (H) by the suspension of bacterial membtane, rat mithochondria and NAD-dependent dehydrogenase were accelerated by paraquat treatment.

Effect of Herbicide Paraquat on Electron Donor and Acceptor (제초제 Paraquat의 전자수용 및 방출에 대한 영향)

  • Kim Mi-Lim;Choi Kyung-Ho
    • Journal of Life Science
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    • v.15 no.2 s.69
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    • pp.311-315
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    • 2005
  • When paraquat was added to the bacterial membrane or mitochondrial suspension, the mixture turned dark blue, but the color was disappeared by aeration. The same phenomenon was seen when electrons were supplied to the paraquat. Blue color appeared from near the cathode, and then spreaded to whole transit system. Coloration was accelerated by addition of alkali, but the color was reduced by addition of acid or oxygen. Paraquat exhibited absorption at ultraviolet region by electron transfer at the concentrations as low as 1.0 mM which did not exert difficulty in showing color reaction. Paraquat caused the increase of the optical density at 340 nm by electron transit, and an aspect of that had a strong resemblance to NADH. The acute toxic action of paraquat seemes to depend on inhibition of energy metabolism cased by paraquat action of electron donor and acceptor.

Growth Inhibition of Newly Emerging Arcobacter butzlrei by Organic Acids and Trisodium Phosphate (새롭게 출현한 Arcobacter butzleri의 유기산과 trisodium phosphate 처리에 의한 생육저해효과)

  • Jang, Jung-Soon;Lee, Young-Duck;Park, Jong-Hyun
    • Korean Journal of Food Science and Technology
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    • v.35 no.6
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    • pp.1169-1173
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    • 2003
  • Growth of a newly emerging pathogen, Arcobacter butzleri, in domestic raw meat was evaluated by various sanitizing agents. One percent of acetic acid, citric acid, lactic acid, and trisodium phosphate (TSP) added to the cell suspension of six A. butzleri strains inhibited their growth within ten minutes, and especially the lactic acid inhibited growth within five minutes. One percent of all the acids at the culture broth inhibited growth completely within one hr. 0.1% of the acids inhibited growth within 72 hr, whereas two percent of TSP had the same effect in one hr. Among the acids, lactic acid had the strongest inhibition activity. Hydrogen peroxide, sodium hypochlorite, and ethanol showed lower inhibiting activities than the above agents. While garlic extract and lactic acid bacteria culture also inhibited A. butzleri, onion extract did not. Therefore, food-borne poisoning of A. butzleri in raw meat could be prevented by organic acid and trisodium phosphate treatments.

Delivery System of Daunorubicin by Red Blood Cells (적혈구를 이용한 Daunorubicin의 배송시스템)

  • Ham, Seong-Ho;Song, Kyung;Ko, Gun-Il;Kim, Jae-Baek;Sohn, Dong-Hwan
    • Journal of Pharmaceutical Investigation
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    • v.24 no.3
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    • pp.131-137
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    • 1994
  • Drug delivery system by the use of red blood cells was established to sustain the release of drugs in the circulatory system by the intravenous injection. The entrapment method by the preswelling technique was re-examined and evaluated for searching the new entrapping conditions without hemolysis. The addition of 4 volume of $0.6{\times}\;hank's$ balanced salt solution (HBSS) into 1 volume of 50% red blood cells suspension did not induce the hemolysis and change the hematocrit level in this experimental condition (within 15 min). Most of daunorubicin could be entrapped into red blood cells within 15 min. While the intracellular adenosine triphosphate (ATP) level followed by the entrapment was reduced to 86% of normal ATP level, the membrane fluidity and the shape factor of red blood cells were not altered. The release rate of daunorubicin from red blood cells was affected by the hemolysis under this condition. To maintain the intracellular ATP in red blood cells, the new reaction buffer was made With the addition of ATP and sodium pyruvate during the entrapment procedure because the hemolysis during the release test would reflect the loss of intracellular ATP that might result in the decrease of the viability in vivo. The addition of ATP raised the intracellular ATP level, which protect the hemolysis during the release test.

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Relationship between Germ Tube Formation, Adherence to Human Buccal Epithelial Cells and Virulence of Candida albicans (Candida albicans의 상피세포에 대한 부착능과 병원성과의 상관관계에 관한 연구)

  • Koh, Choon-Myung
    • The Journal of the Korean Society for Microbiology
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    • v.21 no.4
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    • pp.407-415
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    • 1986
  • This study investigated whether a correlation exists between environmental physical and biochemical factors and adherence of Candida albicans to human buccal epithelial cells by using normal and UV-irradiated strains. The results were as follows: 1. The percentage of germ tube forming activities of normal Candida albicans was 91.5% and UV-irradiated Candida albicans was 15.0%. The $LD_{50}$ of normal strains in mice were $1.0{\times}10\;cells/ml$, but could not be observed in the UV-irradiated strains even with $1.0{\times}10\;cells/ml$. It demonstrated that the virulence is decreased in the UV-irradiated strain. 2. The adherence of normal Candida albicans to human buccal epithelial cells($166{\pm}29{\sim}207{\pm}17\;cells$/100 epithelial cells) was significantly greater than UV-irradiated Candida albicans($99{\pm}21{\sim}131{\pm}25\;cells$/100 epithelial cells). 3. Candida albicans cultured at $37^{\circ}C$ adhered to buccal epithelial cells($166{\pm}16{\sim}207{\pm}17\;cells$/100 epithelial cells) in greater numbers than cultured at $25^{\circ}C$($80{\pm}15{\sim}143{\pm}22\;cells$/100 epithelial cells). 4. On comparison of the adherence of viable and nonviable(heat-killed) Candida albicans to human buccal epithelial cells, the nonviable Candida albicans demonstrated poorer adherence than viable Candida albicans. 5. Adherence in vitro of Candida albicans to human epithelial cells appeared to be effected by the pH. The adherence ability was maximum increased at pH 7.0($187{\pm}22\;cells$/100 epithelial cells) other than experimental pH. 6. The adherence was proportional to the incubation time and the Candida cell concentration in the suspension. 7. A strong correlation was shown between germ tube forming activity and increased adherence of Candida albicans to human epithelial cells, indicating that germ tube forming activity were responsible for candidal virulence.

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Production and Characterization of Nitrate Reductase Deficient Mutants in Petunia parviflora

  • Lee, Cheol-Hee
    • Korean Journal of Plant Resources
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    • v.19 no.6
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    • pp.706-715
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    • 2006
  • Nitrate reductase deficient (NR) mutant lines were selected indirectly by their resistance to 100mM chlorate in cell cultures of P. parviflora. A total of 585 chlorate resistant lines were confirmed by a second passage on a high concentration of chlorate. Frequency of spontaneous mutation was $9.7{\times}10^{-7}$ in 3 month old suspension-cultured cells, and in non-selective media containing amino acids as sole nitrogen source. The frequency of mutation could be increased up to 11-fold by culture for 12 months. Out of 40 randomly selected calli, 22 were fully deficient in NR. The rest of the clones contained a decreased level of NR activity. Further characterization was carried out in 13 mutant lines which were fully deficient in NR and in 5 mutant lines containing residual (0-7.0%) NR activity, as compared to wild-type cells cultured on the same medium. The $NR^-$ mutants were tentatively classified as defective in the NR apoenzyme (nia-type; 11 mutant lines including the 5 with residual NR activity) or in the molybdenum cofactor (cnx-type; 7 mutant lines) by the XDH activity. The cnx-type could be further classified into two groups. In one group (5 mutant lines) of these, the NR activity could be partially restored by nonphysiologically high (1.0mM) molybdate in the culture medium. Both types of $NR^-$ mutants were unable to grow on minimal medium containing nitrate as sole nitrogen source, but grew well on amino acids. They also proved to be extremely sensitive to the standard medium ($MSP_1$) containing nitrate and ammonium. Shoot regeneration was obtained only in the $NR^-$ mutants, which contained residual NR activity, but they so far have failed to grow into plants.

Radiation-induced DNA strand breaks in EL4 cells and mouse spleen lymphocytes (방사선에 의한 EL4 마우스 백혈병세포 및 정상 마우스 비장 임파구 DNA strand breaks의 측정)

  • Kim, Sung-ho;Kim, Tae-hwan;Chung, In-yong;Yoo, Seong-yul;Cho, Chul-koo;Chin, Soo-yil
    • Korean Journal of Veterinary Research
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    • v.31 no.3
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    • pp.329-335
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    • 1991
  • The filter elution technique was used to assay $^{60}Co$ $\gamma$ ray-induced DNA strand breaks(SB) in EL4 mouse leukemia cell and mouse spleen lymphocyte. The lymphocytes were stimulated with lipopolysaccharide (LPS, $20{\mu}g/ml$) to label $[^3H]$ thymidine. EL4 cells and lymphocytes in suspension were exposed at $0^{\circ}C$ to 0Gy, 1Gy, 5Gy, 10Gy or l5Gy for DNA single strand breaks(SSB) assay and 0Gy, 25Gy, 50Gy, 75Gy or 100Gy for DNA double strand breaks(DSB) assay of $^{60}Co$ radiation and elution procedure was performed at pH12.1 and 9.6. The number of DNA strand breaks increased with increasing doses of r rays. The strand scission factor(SSF) was estimated in each experiment (eluted volume 21ml). The slope of SSB EL4 cells was $0.01301{\pm}0.00096Gy^{-1}$ (n=5), the slope of SSB for lymphocytes was $0.01097{\pm}0.00091Gy^{-1}$ (n=5) and the slope of DSB for lymphocytes was $0.001707{\pm}0.0000573Gy^{-1}$ (n=5). Thus EL4 cells were more sensitive to induction of DNA SSB by ionizing radiation than lymphocytes (p<0.005). The ratio of slope of dose-response relationship (SSF versus dose) of lymphocytes DNA SSB as compared with the slope of DNA DSB was 6.4.

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Immunosuppressive effects and pathogenicity of a Korean isolate of reticuloendotheliosis virus in chickens (Reticuloendotheliosis virus의 닭에 대한 면역억제효과와 병원성)

  • Han, Myung-guk;Kim, Sun-joong
    • Korean Journal of Veterinary Research
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    • v.40 no.2
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    • pp.311-323
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    • 2000
  • Immunosuppressive effects of reticuloendotheliosis virus (REV) infection in chickens were investigated. Primary antibody responses to Newcastle disease virus (strain B1) and sheep red blood cells were significantly low in chickens inoculated with the local isolate 89-74 of REV compared to those of uninfected chickens. In chickens infected with REV strain T or 89-74, blastogenesis of spleen cells and peripheral blood lymphocytes (PBL) to concanavalin A (Con A) was severely suppressed. When specific pathogen free (SPF) chickens were inoculated with the isolate, the suppressive effect was observed up to 7 weeks of age while, in the contact infected chickens, the suppression was absent. Similar suppressive effects were observed in chickens inoculated with REV strain T at 2, 3 and 4 weeks of age. When spleen cells or PBL from uninfected chickens were co-cultured with spleen cells or PBL from chickens infected with REV at 1 day-old or 2 week-old, the blastogenesis of the normal cells was suppressed. The suppressive effect of PBL from REV-infected chickens on normal lymphocytes was abrogated by the treatment with trypsin. However the suppressive activity of the REV-infected PBL was not influenced at removing machrophage from the cell suspension by incubation in plastic petri dishes. In addition to the immunosuppression, chickens infected with the REV isolate showed abnormal feather development (nakanuke), anemia, paralysis and retarded growth. Three out of 11 chickens inoculated with the isolate at day-old died between 6 and 9 weeks of age by bacterial infections.

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