• Title/Summary/Keyword: Co-incubation

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Rumen Fermentation was Changed by Feed Inoculation Method in In Vitro (사료 접종 방법에 의한 in vitro 반추위 발효 상성 변화)

  • Yoo, Dae-Kyum;Moon, Joon-Beom;Kim, Han-Been;Yang, Sung-Jae;Park, Joong-Kook;Lee, Se-Young;Seo, Ja-Kyeom
    • ANNALS OF ANIMAL RESOURCE SCIENCES
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    • v.30 no.3
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    • pp.111-120
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    • 2019
  • The objective of this study was to investigate the effect of different feed inoculation method on rumen fermentation in an in vitro. Three experimental treatments were used: control (CON, direct dispersion of feed (2 g) in rumen fluid), combinations of direct dispersion (1 g) and nylon bag (DNB, pore size: 50 ㎛, 1 g), and nylon bag (NB, 2 g). An in vitro fermentation experiment was carried out using strained rumen fluid for 48 h incubation time and timothy was used as a substrate. At the end of the incubation, in vitro dry matter digestibility (IVDMD), in vitro neutral detergent fiber digestibility (IVNDFD), pH, volatile fatty acids (VFA), ammonia nitrogen (NH3-N), and microbial community were evaluated and gas production was estimated at 3, 6, 12, 24, 48 h incubation periods. Gas production was higher in CON than DNB and NB at 6 and 12 h incubation time (p<0.01). There were no differences in final gas production, pH, NH3-N concentration, total VFA production, and VFA profiles among treatments. The IVDMD was lowest in CON (p<0.01) but the IVNDFD was not differed by feed distribution methods. There were no significant differences in general bacteria and fungi. Protozoa count was highest in NB treatment among treatments (p<0.01). The abundance of cellulolytic bacteria, Ruminococcus flavefaciens and Fibrobacter succinogenes, was highest in the CON among treatments (p<0.01).

Effects of Multiple Stress Factors Including Iron Supply on Cell Growth and Lipid Accumulation in Marine Microalga Dunaliella tertiolecta (해양 미세조류 Dunaliella tertiolecta에서 철 공급을 포함한 다중스트레스 인자가 세포성장 및 지질생산에 미치는 영향)

  • Rizwan, Muhammad;Mujtaba, Ghulam;Lee, Kisay
    • Applied Chemistry for Engineering
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    • v.28 no.3
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    • pp.306-312
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    • 2017
  • Changes in the cell growth and lipid accumulation of marine microalga Dunaliella tertiolecta were investigated in response to the combination of different stress factors including the variation of iron supply as a primary stress factor and different options in light irradiation and $CO_2$ supply as a secondary stress factor. High or limited Fe conditions could act as a stress for lipid synthesis. As a secondary stress factor, non-$CO_2$ condition was good for lipid accumulation, but the overall cell growth was sacrificed significantly after a long-time cultivation. Dark condition as a secondary stress factor also favored lipid accumulation and the extent of cell density reduction at the early period in the dark was small compared to other stress conditions. The two-stage cultivation strategy was necessary to maximize lipid production because tendencies of the cell growth and lipid content were not identical under the chosen stress condition. The first stage was for preparing a high cell density under the normal growth-favoring condition and the second stage was the stress condition to induce lipid accumulation in a short time. The short-term (12 h) incubation under the 5X Fe (3.25 mg/L) and dark conditions resulted in the best lipid productivity of 1.44 g/L/d providing 2 g/L inoculum at the second stage.

Impaired Functions of Lymphocytes on Nitric Oxide Production in Endotoxin- Tolerant Mice (내독소내성 마우스에서 Nitric Oxide 생성에 미치는 림프구 부전)

  • Gil, Young-Gi;Kang, Mi-Kyung
    • Journal of Life Science
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    • v.18 no.11
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    • pp.1471-1478
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    • 2008
  • In this study, nitric oxide (NO) production in a macrophage-lymphocyte co-culture system was used to assess the cytokine producing capability of cells during endotoxin tolerance in mice. Incubation of peritoneal macrophages with interferon-$\tau$ (IFN-$\tau$) in the presence of lipopolysaccharide (LPS) augmented NO synthesis. Exogenous tumor necrosis factor-$\alpha$(TNF-$\alpha$) could also replace LPS for the stimulation of NO production. Macrophages co-cultured with splenic lymphocytes showed augmented NO synthesis by LPS alone. However, pretreatment of mice with 2.5 mg/kg LPS completely prevented the lethality and the increase of blood TNF-$\alpha$ and IFN-$\tau$ after the second challenge with a lethal dose of LPS. In addition, when macrophages prepared from LPS-tolerant mice were co-cultured with normal splenocytes, LPS also could not induce the production of NO, even in the presence of exogenous TNF-$\alpha$. Moreover, when normal macrophages were co-cultured with splenocytes obtained from LPS-tolerant mice, stimulation with LPS could not evoke the NO production enhancement. However, this down-regulation was able to reverse by exogenous IFN-$\tau$ or concanavalin A (ConA), a stimulator of IFN-$\tau$ production. Our results indicate that not only macrophages but also lymphocytes contribute to LPS tolerance. As INF-$\tau$ can enhance the expression of TNF-$\alpha$, the decrease of INF-$\tau$synthesis from lymphocytes may orchestrate with the decrease of TNF-$\alpha$ synthesis from LPS-tolerant macrophages for the production of tolerant state and the prevention of excessive inflammation. Therefore, LPS tolerance may be exploited for prophylaxis of severe sepsis in patients at risk.

Screening of lovastatin-producing strains by PCR using lovastatin biosynthesis genes (Lovastatin 생합성 유전자를 이용한 lovastatin 생산균주의 탐색)

  • Ko, Hee-Sun;Kim, Hyun-Soo
    • KSBB Journal
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    • v.24 no.2
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    • pp.163-169
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    • 2009
  • Lovastatin (also known as Mevinolin, Mevacor, and Monacolin K), an inhibitor of the HMG-CoA reductase produced by Aspergillus terreus and other fungi, is used to reduce serum cholesterol levels in human beings. It is derived biosynthetically from two polyketides. One of these is a nonaketide that undergoes cyclization at a hexahydronaphthalene ring system, and the other is a simple diketide, 2-methylbutyrate. Two primer pairs were designed based on the amino acid sequences of lovastatin polyketide synthase and lovastatin diketide synthase for the PCR screening of lovastatin-producing strains. Among the seven selected strains, SJ-2 evidenced the highest level of lovastatin production in both liquid and solid cultures. Soybeans with SJ-2 were treated via 1 hour of heat shock at $30^{\circ}C$ for the mass production of lovastatin. The heat-treated soybeans were inoculated on rice bran and the koji extract was obtained after 15 days of incubation. It yielded the highest level of lovastatin production among the strains, and also evidenced 75% inhibition activity against HMG-CoA reductase. We developed an efficient PCR screening method for lovastatin-producing strains, using lovastatin biosynthesis genes.

Comparison between Percoll and Sil-Select Methods on the Human Spermatozoa Treatment (인간정자의 처리에 있어서 Percoll과 Sil-Select 방법의 비교)

  • Moon, Shin-Yong;Ryu, Buom-Yong;Shin, Hyun-Ah;Oh, Sun-Kyung;Suh, Chang-Suk;Kim, Seok-Hyun;Choi, Young-Min;Kim, Jung-Gu;Choi, Kyu-Hong;Lee, Jin-Yong
    • Clinical and Experimental Reproductive Medicine
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    • v.27 no.4
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    • pp.339-344
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    • 2000
  • Objective: To evaluate silane-coated silica particles (Sil-select) as an alternative to polyvinylpyrrolidone-coated particles (Percoll) for gradient separation of spermatozoa, for use in assisted reproduction. Methods: 20 normal semen based on WHO criteria were included in this study. Recovery of motile and morphologically normal spermatozoa after using two-layer Percoll and Sil-select gradient respectively was recorded. Motility, HOST (hypoosmotic swelling test) and the detection of malondialdehyde for LPO (lipid peroxidation) after 24 h of incubation at $37^{\circ}C$ in a 5% $CO_2$ incubator were compared. Results: Percoll (78.5%) and Sil-select (79.1%) showed a significant increase in the motility compared to ejaculate (60.9%) but no difference between Percoll and Sil-select. Normal sperm morphology significantly increased after Percoll (57.6%) and Sil-select (53.7%) compared to ejaculate (35.8%) but no difference between Percoll and Sil-select. No differences in the recovery of motile spermatozoa and motility, HOST and the production of malondialdehyde after 24 h incubation were found when comparing the use of Percoll and Sil-select. Conclusion: Sil-select seems to be an attractive alternative to Percoll for sperm separation in assisted reproduction.

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Establishment of Efficient Microinjection System in the Porcine Embryos

  • Malaweera, Don Buddika Oshadi;Ramachandra, Sisitha;Wu, Jun-Bo;Oh, Seung-Kyu;Kim, Seung-Hwan;Kim, Seok-Joong;Shin, Sang-Tae;Cho, Jong-Ki
    • Journal of Embryo Transfer
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    • v.29 no.1
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    • pp.59-66
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    • 2014
  • Transcription activator like effector nucleases (TALENs) are artificial restriction enzymes generated by fusing a TALE DNA binding domain to a DNA cleavage domain which remove and introduce specific genes to produce transgenic animals. To investigate the efficient laboratory techniques for the injection of TALEN mRNA, pEGFP-N1 commercial plasmid were microinjected into porcine parthenogenetic and in vitro fertilization (IVF). In Experiment 1, to investigate injection time, compared 4 different time durations (2 hr, 4 hrs, 6 hrs & 8 hrs) after post activation of parthenogenetic embryos and after 6 hrs of co-incubation with sperms in IVF embryos. There were significant difference (P<0.05) in development to the blastocysts (4.4, 8.9, 3.9, 0.6%), GFP expression in blastocysts (1.3, 5.7, 2.3, 0.0%) which injected after post activation of 4 hrs compared with other 3 groups. IVF embryos after 2 hrs and 4 hrs injected were expressed GFP significantly higher than rest of two groups (P<0.05). In Experiment 2, compared development of 2 different concentrations ($20ng/{\mu}l$ and $50ng/{\mu}l$) of EGFP injection. There were significant difference (P<0.05) between two treatments which has higher cleavage (58.8 vs 41.9%), blastocysts development rate (13.0 vs 11.1%) and GFP expressed blastocysts (5.7 vs 0.0%) in $20ng/{\mu}l$ than the $50ng/{\mu}l$ in parthenogenetic embryos. In IVF embryos, only $20ng/{\mu}l$ injected embryos were expressed GFP (4.2%) after 7 days of incubation and 77.3 vs 64.7% of cleavage, 26.4 vs 23.5% development to blastocysts. In Experiment 3, three different volumes (5, 10 and 20 pl) were microinjected into porcine embryos to determine the most appropriate volume. Out of 3 groups, significantly higher development rates of cleavage (68.3, 58.0, 29.3%), blastocysts (11.7, 12.7, 0.5%) and GFP expressed blastocysts (2.9, 7.8, 0.0%) were shown in the 10 pl group (P<0.05). In conclusion, these results imply that $20ng/{\mu}l$ concentration, 10 pl of volume and injection at 4 hrs after post activation for parthenogenetic and 2~4 hrs after IVF, $20ng/{\mu}l$ concentration and 10 pl volume for IVF embryos were more effective microinjection conditions.

Effect of Gamma Ray Irradiation on the Availability of Applied $^{65}Zn$ to Applied at Organic Matter Added Soils under Laboratory Conditions

  • Kim, Tai-Soon;Han, Kang-Wan
    • Nuclear Engineering and Technology
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    • v.11 no.2
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    • pp.105-110
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    • 1979
  • In order to evaluate the effect of gamma ray irradiation to the soils added with several organic matters on release sad fixation of $^{65}Zn$, a soil incubation test was carried out by use of an acidic and a calcareous soil. The results obtained were summarized as follows : DTPA extractable $^{65}Zn$ decreased gradually with elapsed time of incubation owing to fixation of $^{65}$Zn by the soils but after four weeks DTPA extractable $^{65}Zn$ was reached an equilibrium state. The most parts of $^{65}Zn$ activity in the equilibrium state was found in the solid phase. The extent of zinc fixation in acidic Keumgok and calcareous Jecheon soils represented as activity basis was 89 and 93.7 per cent of applied $^{65}Zn$ respectively. Increased tendency of DTPA extractable zinc was shown by irradiating gamma ray to both soils because of $^{65}Zn$ fixation could be decreased by irradiation. The extractability of fixed $^{65}Zn$ by DTPA solution was 38.9 per cent in acidic Keumgok soil and only 9.5 per cent in calcareous Jecheon soil. Release of fixed $^{65}Zn$ by irradiation of gamma ray is closely related to pH of both soils. Much more $^{65}Zn$ could be released under low pH condition. Effect of gamma ray irradiation to organic matter sources on release of $^{65}Zn$ could not be differentiated. But in the case of nonirradiated soils, however, glucose treated soils couldrelease much more the extractable zinc as compared to those of the straw or cellulose treatment.

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Synthesis and Evaluation of F-18 Labeled 2'-Deoxy-2'-fluoro-5-methyl-1-β-L-arabinofuranosyluracil (L-[18F]FMAU)

  • Jo, Nam-Hyun;Moon, Byung-Seok;Hong, Su-Hee;An, Gwang-Il;Choi, Tae-Hyun;Cheon, Gi-Jeong;Cho, Jung-Hyuck;Yoo, Kyung-Ho;Lee, Kyo-Chul;Oh, Chang-Hyun
    • Bulletin of the Korean Chemical Society
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    • v.28 no.12
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    • pp.2449-2453
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    • 2007
  • L-[18F]FMAU ([18F]1b) was prepared from the precursor 2-O-[(trifluoromethyl)-sulfonyl]-1,3,5-tri-Obenzoyl- α-L-ribofuranose, by coupling the radioactive fluoro-sugar with the corresponding silylated thymine in 4 steps. The final products, including the α and β anomers, were purified using reverse phase HPLC with an appropriate solvent (5% CH3CN/H2O) at a flow rate of 3.0 mL/min. The total elapsed time of synthesis was about 180-200 min from EOB. The α/β anomeric ratio of the compounds was about 1:9, and the radiochemical purity of the product (β-form) was >98% with decay-corrected yields of 25-35%. All radioactive samples were confirmed using co-injection with pure non-radioactive analogues in every step. In the cellular uptake in vitro test of herpes simplex virus-thymidine kinase (HSV1-TK) gene expressed cells, the percent uptake of injected dose (%ID) of L- and D-FMAU was 37.28 and 65.86, respectively after 240 min incubation. However, the relative uptake (MCA-TK/MCA cellular uptake ratio) of L-FMAU was higher than that of D-FMAU (%ID of L-FMAU, 0.36 and D-FMAU, 0.93 after 240 min incubation in MCA cells). This means that L-FMAU will show better specific HSV1-TK gene expressed cell uptake for selective HSV1-TK gene imaging.

Simultaneous Determination of Chromium (III) and Chromium(VI) by High Performance Liquid Chromatography(HPLC) (고성능 액체크로마토그래피(HPLC)를 이용한 3가, 6가 크롬의 동시정량에 관한 연구)

  • Roh, Jae Hoon;Kim, Chi Nyon;Kim, Choon Sung;Kim, Kyoo Sang
    • Journal of Korean Society of Occupational and Environmental Hygiene
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    • v.4 no.2
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    • pp.189-197
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    • 1994
  • Analytic methods for Cr(VI) level in industrial hygienic field were suggested by the National Institute for Occupational Safety and Health(NIOSH method 7600, 7604). There were growing needs for measurement of Cr(III) and Cr(VI) levels simultaneously. Two analytical methods were suggested to determine Cr(III) and Cr(VI) levels simultaneously. The one is method by using reversed phase high peformance liquid chromatography(HPLC) and the other is by using ion exchange HPLC. The purpose of this work was to evaluate the usefulness of these two analytic methods. For the difference of ionic charges of Cr(III)-ethylendiamine tetraacetic acid(EDTA) chelate and $CrO_4{^-2}$, we could detect them simultaneously by ion exchange HPLC. Also, we attempted to determine the levels of Cr(III) and Cr(VI) chelated with sodium diethyldithiocarbamate(NaDDTC) by using reversed phase HPLC. The confirmation of Cr(III) and Cr(VI) were checked by fraction collector and nameless atomic absorption spectrometer. The optimal conditions for the formation of Cr(III)-EDTA chelate were two hours incubation period with pH 5. Cr(III)-EDTA and Cr(VI) in EDTA solution were successfully separated by anion exchange column using $Na_2CO_3/NaOH$ mixture as mobile phase. Peaks of Cr(III)-EDTA and Cr(VI) in EDTA were identified at 5 minutes and 7 minutes of retention time respectively by the ion exchange HPLC. The formation of Cr(III)-NaDDTC and Cr(VI)-NaDDTC chelates were twelve hours incubation period. Cr(III)-NaDDTC and Cr(VI)-NaDDTC chelates were separated by reversed phase column using methanol and water mixture as mobile phase. Peaks of Cr(VI)NaDDTC and Cr(III)-NaDDTC chelates were identified at 13 minutes and 26 minutes of retention time respectively by the reversed phase HPLC. Due to reduction of Cr(VI) to Cr(III), it seems to be not suitable for simultaneous determination of Cr(III)-NaDDTC and Cr(VI)-NaDDTC chelates by reversed phase HPLS. Simultaneos determination of Cr(III) and Cr(VI) by ion exchange HPLC was more accurate and simple method.

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Effects of the Medicinal Plants Against Streptococcus Sobrinus B13 in Hexane Fraction (수종의 생약 헥산분획 추출물에 의한 Streptococcus sobrinus 억제 효과)

  • Youn, Hye-Jeong;Ha, Myung-Ok;Lee, Kyung-Hee;Park, Young-Nam;Cho, Min-Jeong
    • Journal of dental hygiene science
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    • v.7 no.3
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    • pp.197-200
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    • 2007
  • This study was carried out to assess anticariogenic activity from natural products. The hexane extracts of 11 medicinal plants were tested for the antimicrobial activities against a cariogenic bacterium, S. sobrinus B13. The hexane extracts of 11 medicinal plants such as Alisma orientale, Cyperus rotundus, Saussurea lappa, Atractylodes japonica, Kalopanax pictus, Angelica gigas, Salvia miltorrhiza, Lonicera japonica, Gardenia jasmoides, Aurantii nobilis percarpium and Bupleurum lorgiradiatum were used for this study, and S. sobrinus B13 was cultured with 0.5% extracts of the medicinal plants, in trypticase soy broth medium under 5% $CO_2$ for 0, 6, 12, 24 hours at $37^{\circ}C$. The obtained results were as follows. Immediately after culturing for inoculation, Kalopanax pictus and Cyperus rotundus showed better antimicrobial activities than those of others. After culturing for 6 hours, the order of effective antimicrobial activity was Salvia miltorrhiza, Angelica gigas. After incubation for 12 hours, Salvia miltorrhiza showed the best antimicrobial activity. After incubation for 24 hours Salvia miltorrhiza, Cyperus rotundus, Saussurea lappa, Atractylodes japonica had better antimicrobial activity than others. In conclusion, among Hexane extracts of 11 medicinal plants, Cyperus rotundus, Saussurea lappa, Atractylodes japonica showed effective antimicrobial activity.

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