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Ceriporia sp. ZLY-2010 in Biodegradation of Polychlorinated Biphenyls : Extracellular Enzymes Production and Effects of Cytochrome P450 Monooxygenase (Ceriporia sp. ZLY-2010에 의한 폴리염화비페닐류의 생분해 : 균체 외 효소활성 및 cytochrome P450 monooxygenase 영향)

  • Hong, Chang-Young;Gwak, Ki-Seob;Lee, Su-Yeon;Kim, Seon-Hong;Jeong, Han-Seob;Choi, In-Gyu
    • Journal of the Korean Wood Science and Technology
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    • v.39 no.6
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    • pp.469-480
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    • 2011
  • In this study, to determine the ligninase activity related to the PCBs degradation of Ceriporia sp. ZLY-2010, the protein contents and manganese peroxidase (MnP) and laccase activities during cultivation on shallow stationary culture (SSC) medium were observed. 4 PCB congeners were selected depending on the number of chlorine substituted on biphenyl. Furthermore, to examine the effects of cytochrome P450 monooxygenase, the inhibition of cytochrome P450 monooxygenase was evaluated by measuring the biodegradation rate when inhibitor such as 1-aminobenzotriazole was added. The extracellular protein contents were affected by PCB congeners in culture media. The total protein in the culture medium showed the biggest differences between the samples containing 2,2',4,4',5,5'-hexachlorobiphenyl and the control. On the other hand, MnP and laccase activity showed dominant increases within samples containing 4,4'-dichlorobiphenyl and 2,3',4',5-tetrachlorobiphenyl. Cytochrome P450 monooxygenase was inhibited by adding inhibitor, 1-aminobenzotriazole in low concentration. Only 2.73% of 2,3',4',5-tetrachlorobiphenyl was degraed on day 1, and biodegradation of 2,2',4,4',5,5'-hexachlorobiphenyl was inhibited as well, showing about 20% of biodegradation rate.

A Fish Nodavirus Isolated from Cultured Sevenband Groupe, Epinephelus septemfasciatus (양식 능성어로 부터 Fish Nodavirus 분리)

  • Sohn, Sang-Gyu;Park, Myoung-Ae;Oh, Myung-Joo;Chun, Seh-Kyu
    • Journal of fish pathology
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    • v.11 no.2
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    • pp.97-104
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    • 1998
  • Since 1989, mass mortality has repeatly occurred in cage-cultured sevenband grouper, Epinephelus septemfasciatus along the southern coast of Korea in the summer season and usually reached over 80% within a few months. Diseased fish showed the clinical signs of anorexia, dark coloration, loss of eqilibrium, spinal swimming behaviour, vertebral deformity and inflation of swim bladder. Histopathologically, necrosis and/or vacuolation of the nerve cells in the brain and retina were observed. We previously reported that the causative agent was filtrable. The causative agent was not culturable in various fish cells; RTG-2, CHSE-214, BF-2, EPC and FHM. However, electron microscopic observation revealed unenveloped icosahedral viral particles with about 30 nm in diameter in the cytoplasm of nerve cells of the brain. The characteristics of the virus was tested by an artificial infection with the filtrate of the homogenate of diseased fish. The pathogenicity of the virus was retained after treatment with ether or heat ($50^{\circ}C$, 30 min) but partly lost by pH 3 or 11 treatment. These results suggest that the causative agent are similar to the fish nodavirus. In order to compare the causative agent with a fish nodavirus, Striped Jack Nervous Necrosis Virus (SJNNV), a polymerase chain reaction (PCR) was performed with primers specific to SJNNV. As a result, about 430 by PCR products were detected from the brain and the eye of both naturally and artificially infected sevenband grouper. All these results represent that the mass mortality in the cultured sevenband grouper is caused by the infection of a nodavirus similar to SJNNV and this is the first report of a fish nodavirus from the cultured sevenband grouper in Korea.

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A New Rhabdovirus (HRV-like) Isolated in Korea from Cultured Japanese Flounder Paralichthys olivaceus (양식산 넙치로부터 HRV-like Rhabdovirus의 분리)

  • Oh, Myung-Joo;Choi, Tae-Jin
    • Journal of fish pathology
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    • v.11 no.2
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    • pp.129-136
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    • 1998
  • In March 1997, a new rhabdovirus was isolated from moribund cultured Japanese flounder Paralichthys olivaceus in sea water tank and cage culture systems in Kyung-Nam and Chun-Nam province, Korea. At temperature $15^{\circ}C$ the virus replicated and induced cytopathic effects (CPE), which progressed to eventual cytolysis, in susceptible cell lines, including RTG-2 and EPC. The CHES-214 cell line was refractory. Virus particles were bullet-shaped and measured $70nm{\times}100$ to 150 nm in size. The isolate was sensitive to pH 3, to diethyl ether, and to heat ($50^{\circ}C$ 5 min, $60^{\circ}C$ 1 min). Viral replication was not inhibited by $10^{-4}$ M 5-iododeoxyuridine. Virus infectivity was reduced by anti-HRV (8401-H) rabbit serum, but can not reduced by antisera against infectious hematopoietic necrosis virus (IHNV), chum salmon reovirus (CSV), retrovirus of salmonid (RVS) and infectious pancreatic necrosis virus (IPNV). HRV virus antigen was detected by fluorescent antibody test (FAT) in the cytoplasm of infected EPC cell. Purified isolates virions were composed of: polymerase (L), glycoprotein (G), nucleoprotein (N) and 2 matrix proteins (M1 and M2). Based upon their relative mobilities, the estimated molecular weights of the proteins were: L, 160 kDa; G, 55 kDa; N, 45 kDa; M1, 26 kDa; and M2, 22 kDa.

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Purification and Gene Analysis of Peptidyl Prolyl cia-trans Isomerase from Bacillus stearothermophilus (Bacillus stearothermophilus Peptidyl Prolyl cis-trans Isomerase의 정제 및 유전자 분석)

  • 김동주
    • The Korean Journal of Food And Nutrition
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    • v.15 no.2
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    • pp.104-111
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    • 2002
  • The peptidyl prolyl sis-trans isomerase (PPIase, EC 5.2.1.8) from bacillus stearothermophilus was extracted from the cells treated with by lysozyme. PPIase was purified from the cell extracts by heat treatment, ammonium sulfate precipitation, ion exchange chromatography and finally gel filtration, sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). The molecular weight of the purified PPIase was estimated as 18kDa by SDS-PAGE. The 39 amino acid residues from the N-terminus were determined by the protein sequencer. The enzyme showed the optimum pH at 8.0 and was stable at the range of pH 7.0∼8.0. The enzyme was considerably stable after heat treatment at 60$\^{C}$ for 30minutes, and the enzyme was quite stable up to 65$\^{C}$. The presence of the PPIase in the refolding solution accelerated the isomerization rate of the assay peptide. PPIase gene of Bacillus stearothermophilus was screened from a genomic library by plaque hybridization using the A-l primer as a probe. A PPIase positive plaque contained a 3.0kb insert of the chromosomal DNA. A 3.0kb fragment was subcloned into pUC18, resulting pPI-40. A DNA fragment encoding the N-terminal portion of the PPIase in pPI-40 was amplified by polymerase chain reaction(PCR) method using the A-1 and B-2 primers. The amplified fragment was cloned into the Sma I site of pUC18 and recombinant plasmid was designated as pSN-18. The nucleotide sequence of 167bp fragment was determined. The deduced amino acid sequence of PPIase was completely matched with the determined N-terminal amino acid sequence of PPIase B. stearothermophilus.

Characterization of Extracellular Cholesterol Oxidase Produced from Soil Microorganism (토양 미생물로부터 생산된 Extracellular Cholesterol Oxidase의 특성)

  • Park, Jeong-Su;Jeong, Jong-Moon
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.37 no.11
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    • pp.1507-1514
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    • 2008
  • Cholesterol oxidase catalyses the conversion of cholesterol to 4-cholesten-3-one. This enzyme has been used for clinical assay of human serum cholesterol and for reduction of cholesterol level in foods and feeds. In order to search the microorganism which has a high extracellular and stable activity of cholesterol oxidase, soil microorganisms were screened. As a result, the one with the highest extracellular cholesterol oxidase activity was obtained and named as the BEN 115. The BEN 115 strain was identified as one of the Nocardia species based on our taxonomic studies. The cholesterol oxidase from this strain was shown to have two bands of extracellular proteins on SDS-PAGE and Western blot. Their molecular masses were estimated to be about 55 and 57 kDa, respectively. In addition, this cholesterol oxidase was considerably stable at the broad range of pH $3.5{\sim}9.5$ and at the temperature of $25{\sim}55^{\circ}C$. The optimum pH and temperature of this cholesterol oxidase were pH 5.5 and $35^{\circ}C$, respectively. The activity of extracellular cholesterol oxidase could be enhanced 1.6 to 2.0 folds by the addition of nonionic detergent such as Triton X-114, Triton X-100, or Tween-80 into the culturing broth. The substrate specificities against campesterol, sitosterol and stigmasterol were measured to be 50%, 50%, and 27%, respectively, compared to the cholesterol. These results suggest that Nocardia sp. BEN 115 may be useful as a microbial source of cholesterol oxidase production.

Effects of Artemisia Capillaris Thunberg on Apoptosis in HeLa Cells (사철쑥의 HeLa 세포고사 효과)

  • Lee, Hyoung-Ja;Kim, Kee-Hwan;Park, Jong-Kun;Hwang, Eun-Hee
    • Journal of Nutrition and Health
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    • v.41 no.1
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    • pp.22-30
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    • 2008
  • This study analyzes the apoptosis of HeLa cells to see if we can use the Artemisia capillaris Thunberg for the prevention of chronic degenerative diseases. We used the HeLa cells to see what effects the A. capillaris Thunberg had on apoptosis of the cancer cells. We checked the cell activity, cell morphological change, DNA fragmentation, and DNA content after administering 0, 100, 500, 1000, and $2000{\mu}g/ml$ methanol, ethyl acetate, n-butanol extract of the A. capillaris Thunberg. As for the cell viability, the increase of concentration of methanol and ethyl acetate decreased the survival rate of the cell, but the phenomenon was much weakened in n-butanol extract and was not observed in aqueous extract. The higher the density of the methanol, ethyl acetate, n-butanol and aqueous extract was, the lower the survival rate of the HeLa cell was. These extracts obstructed the cell cohesion and caused the blebbing of he cell membrane and fragmentation of the nucleus, both of which are symptoms of apoptosis. Laddering-pattern DNA fragmentation was observed in the groups that were treated with the $1000{\mu}g/ml$ and $2000{\mu}g/ml$ of methanol extract. The DNA content of the cells apoptosis measured by fluorescent-activated cell sorter (FACS) increased as the density of the methanol, ethyl acetate and butanol extract increased. The result of the study shows that A. capillaris Thunberg fosters the apoptosis of HeLa cells, which suggests that the A. capillaris Thunberg has a great potential value as food additives, medicinal supplements for patients with chronic diseases, and preventive measures against cancer.

Cellular Responses of the TNT-degrading Bacterium, Stenotrophomonas sp. OK-5 to Explosive 2,4,6-Trinitrotoluene (TNT) (폭약 2,4,6-Trinitrotoluene에 노출된 분해세균 Stenotrophomonas sp. OK-5의 세포반응)

  • 장효원;송승열;김승일;강형일;오계헌*
    • Korean Journal of Microbiology
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    • v.38 no.4
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    • pp.247-253
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    • 2002
  • The cellular responses of TNT-degrading bacterium, Stenotrophomonas sp. OK-5 to explosive 2,4,6-trini-trotoluene (TNT) as an environmental contaminant were examined. Survival of the strain OK-5 with time in the presence of different concentrations of TNT under sublethal conditions was monitored, and viable counts paralleled the production of the stress shock proteins in this bacterium. Total cellular fatty acids analysis showed that strain OK-5 produced or disappeared several different kinds of lipids when grown on TNT media than when grown on TSA. Under scanning electron microscope, the cells treated with 0.5 mM TNT for 12 hrs showed irregular rod shapes with wrinkled surfaces. Analyses of SDS-PAGE and Western blot using anti-DnaK and anti-GroEL revealed that several stress shock proteins including 70 kDa DnaK and 60 kDa GroEL in strain OK-5 were newly synthesized at different TNT concentrations in exponentially growing cultures. 2-D PAGE of soluble protein fractions from the culture of OK-5 exposed to TNT demonstrated that approximately 300 spots were observed on the silver stained gel ranging from pH 3 to pH 10. Among them, 10 spots significantly induced and expressed in response to TNT were selected and analyzed. As the result of internal amino acid sequencing with ESI-Q TOF, two proteins, spot #1 and spot #10 were assigned the DnaK protein XF2340 of Xylella fastidiosa and stress-induced protein of Mesorhizobium loti, respectively.

Isolation and Identification of Halotolerant Bacillus sp. SJ-10 and Characterization of Its Extracellular Protease (세포외 Protease를 생산하는 내염성 Bacillus sp. SJ-10 균주의 분리 동정 및 효소 특성)

  • Kim, Eun-Young;Kim, Dong-Gyun;Kim, Yu-Ri;Choi, Sun-Young;Kong, In-Soo
    • Korean Journal of Microbiology
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    • v.45 no.2
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    • pp.193-199
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    • 2009
  • A bacterium producing the halotolerant extracellular protease was isolated from squid jeotgal, and was identified as Bacillus sp. SJ-10 based on morphological, physiological and biochemical characteristics, as well as phylogenetic analysis using 16S rRNA gene sequence. The strain grew at $20^{\circ}C\sim55^{\circ}C$, pH 5~8, and 0%~14% NaCl and optimal growth conditions were $35{\pm}5^{\circ}C$, pH 7, and 5% NaCl. The major cellular fatty acids were anteiso-$C_{15:0}$, anteiso-$C_{17:0}$, and $C_{16:0}$ DNA G+C content was 50.58 mol% and menaquinone consisted of MK-7 Phylogenic analysis based on the 16S rRNA gene sequence indicated that SJ-10T belongs to the genus Bacillus. About 40 kDa of the salt-tolerant protease was purified by 40% ammonium sulfate saturation and Mono Q column chromatography. The optimal activity of the protease was pH 8 and stable at pH 5~10. The optimum temperature and NaCl concentration were $35{\pm}5^{\circ}C$ and $5{\pm}1%$, respectively.

Dissolved Oxygen at the Bottom Boundary Layer of the Ulleung Basin, East Sea (동해 울릉분지 해저 경계면의 용존산소)

  • Kang, Dong-Jin;Kim, Yun-Bae;Kim, Kyung-Ryul
    • Ocean and Polar Research
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    • v.32 no.4
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    • pp.439-448
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    • 2010
  • General consensus on typical vertical profile of dissolved oxygen in the Ulleung Basin is that dissolved oxygen concentration beyond 300 m decreases with increasing depth. However, the results of our observations in 2005 and 2006 revealed three different dissolved oxygen distribution types in the deep layer of the Ulleung Basin. The first type showed oxygen concentration decreasing with increasing depth (Type-1), the second showed oxygen concentration decreasing very sharply near the bottom boundary layer but constant in the bottom adiabatic layer (Type-2), the final was of the oxygen minimum layer above the bottom boundary layer (Type-3). Type-2 was the most common pattern in the Ulleung Basin. Type-1 was most common close to the Japan Basin, including the Ulleung Interplane Gap, while Type-3 was found around Dok do. Oxygen Consumption Rate (OCR) at surface sediment estimated using the dissolved oxygen distribution at the bottom boundary layer was $0.2{\sim}5.8\;mmol{\cdot}m^{-2}d^{-1}$, which coincided with OCR from direct sediment incubation. This implies that organic matter decomposition at surface sediment may play an important role in dissolved oxygen distribution patterns at the bottom boundary layer of the Ulleung Basin.

Conservation Environment for Mural Tomb in Goa-ri, Goryeong (고령 고아리 벽화고분의 보존환경 연구)

  • Jeong, Seon Hye;Lee, Hyun Ju;Lee, Min Young;Chung, Yong Jae
    • Journal of Conservation Science
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    • v.33 no.3
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    • pp.189-201
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    • 2017
  • This study focused on the Gaya mural tomb located in Goa-ri, Goryeong-gun, North Gyeongsang Province, Republic of Korea. Environmental factors such as ambient temperature, relative humidity and surface temperature, and microbial distribution on mural tombs were monitored for one year to gather data on the conservation environment of the mural tombs. The average internal temperature difference with reference to the ambient outer temperature was observed to be $11.7^{\circ}C$ for the monitoring period and the internal temperature of the tomb was found to change periodically every one or two months in response to the outer temperature. The highest temperature was observed in September and the lowest in March. The relative humidity in the mural tomb remained constant at 100%. Between December and April, condensation occurred on the ceiling of the main room of the tomb, where the murals are located. On the ceiling of the aisle, the condensation occurred throughout the year. The inside and surface wall were isolated from microorganisms, which could grow when a suitable growth environment suitable is established. Based on microbial growth temperature conditions, risk periods of microbial hazards were established, where in the period from August to October was identified as the most dangerous.