• Journal of Microbiology and Biotechnology
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• 제26권1호
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• pp.160-170
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• 2016

The increasing spread of antibiotic-resistant pathogens has raised the interest in alternative antimicrobial treatments. In our study, the functionally active gram-negative bacterium bacteriophage CP933 endolysin was produced in Nicotiana benthamiana plants by a combination of transient expression and vacuole targeting strategies, and its antimicrobial activity was investigated. Expression of the cp933 gene in E. coli led to growth inhibition and lysis of the host cells or production of trace amounts of CP933. Cytoplasmic expression of the cp933 gene in plants using Potato virus X-based transient expression vectors (pP2C2S and pGR107) resulted in death of the apical portion of experimental plants. To protect plants against the toxic effects of the CP933 protein, the cp933 coding region was fused at its Nterminus to an N-terminal signal peptide from the potato proteinase inhibitor I to direct CP933 to the delta-type vacuoles. Plants producing the CP933 fusion protein did not exhibit the severe toxic effects seen with the unfused protein and the level of expression was 0.16 mg/g of plant tissue. Antimicrobial assays revealed that, in contrast to gram-negative bacterium E. coli (BL21(DE3)), the gram-positive plant pathogenic bacterium Clavibacter michiganensis was more susceptible to the plant-produced CP933, showing 18% growth inhibition. The results of our experiments demonstrate that the combination of transient expression and protein targeting to the delta vacuoles is a promising approach to produce functionally active proteins that exhibit toxicity when expressed in plant cells.

• The Korean Journal of Physiology and Pharmacology
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• 제24권1호
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• pp.101-110
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• 2020

Transient receptor potential canonical 4 (TRPC4) channel is a nonselective calcium-permeable cation channels. In intestinal smooth muscle cells, TRPC4 currents contribute more than 80% to muscarinic cationic current (mIcat). With its inward-rectifying current-voltage relationship and high calcium permeability, TRPC4 channels permit calcium influx once the channel is opened by muscarinic receptor stimulation. Polyamines are known to inhibit nonselective cation channels that mediate the generation of mIcat. Moreover, it is reported that TRPC4 channels are blocked by the intracellular spermine through electrostatic interaction with glutamate residues (E728, E729). Here, we investigated the correlation between the magnitude of channel inactivation by spermine and the magnitude of channel conductance. We also found additional spermine binding sites in TRPC4. We evaluated channel activity with electrophysiological recordings and revalidated structural significance based on Cryo-EM structure, which was resolved recently. We found that there is no correlation between magnitude of inhibitory action of spermine and magnitude of maximum current of the channel. In intracellular region, TRPC4 attracts spermine at channel periphery by reducing access resistance, and acidic residues contribute to blocking action of intracellular spermine; channel periphery, E649; cytosolic space, D629, D649, and E687.

• 한국분무공학회지
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• 제15권2호
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• pp.74-82
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• 2010

In order to understand the behavior of transient gaseous injection used in an LPG (Liquefied Petroleum Gas) engine, turbulent incompressible transient jets with butane and propane were measured and analyzed at pressures of 1.5 bar and 2.0 bar with injector diameters of 3 mm and 5 mm. Mie-scattering method with a tracer was used, and images were processed to investigate the behavior of butane and propane jets. Distances from the nozzle to transition region were measured as $L_e/d_{inj}$=4.35~19.4, where $L_e$ and $d_{inj}$ indicate respectively a distance from nozzle to transition point and nozzle diameter. Slits and tubes around jet at near-field were introduced to measure the effect of entrainment and the diameter of jet, which revealed that the entrainment of surrounding air is significant for developing jet diameter. When the entrainment is restricted, the behavior of jet became deviating from the baseline. It was found that the virtual origin located outside of a nozzle towards jet tip within the conditions of this work, and its location was estimated as $x_o/d_{inj}$=0.56~7.25, where $x_o$ is a distance from nozzle to virtual origin.

• 동의생리병리학회지
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• 제16권3호
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• pp.594-601
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• 2002

The current study was carried out to evaluate neuroprotective effects of Citri Pericarpium after transient global ischemia in gerbils. Male Mongolian gerbils weighing 60-80g were anesthetized with 2% isoflurane mixed with 30% oxygen and 70 % nitrogen. Bilateral common carotid arteries were occluded for 5 minute with microaneurysm dips. On 3 or 7 days after ischemic surgery, the gerbils were sacrificed. The brain were removed, embedded in paraffin and sectioned at 8㎛-thickness. Gerbils that received ischemic insult for 5 min showed extensive neuronal damage in the hippocampal CA1 region, and the number of viable neuronal cell was 51.0±2.5/mm, 32.2% of normal group at 7 days after ischemic surgery. In animals that underwent the extract of Citri Pericarpium treatment, the number of viable neuronal cell were significantly better preserved at 110.58±3.58/mm, 72.0% of normal group than those of ischemic group (P<0.01). In the immunohistochemistry of Bax and Bcl-2, the Citri Pericarpium treated group down-regulated the expression of Bax protein at 72hr after transient global ischemia. In contrast, Bcl-2 protein level was not changed. The appearance in TUNEL assay is similar to the pattern of Bax protein. The water extract of Citri Pericarpium significantly reduced the number of TUNEL-positive CA1 pyramidal neurons at 72hr. The results suggest that Citri Pericarpium has potential neuroprotective effects in the transient global ischemia and the increase in the ratio of Bcl-2 to Bax may contribute to the anti-apoptotic effect of Citri Pericarpium.

• 한국환경농학회지
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• 제40권3호
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• pp.186-193
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• 2021

BACKGROUND: Before generating transgenic plant using the CRISPR-Cas9 system, the efficiency test of sgRNAs is recommended to reduce the time and effort for plant transformation and regeneration process. The efficiency of the sgRNA can be measured through the transient expression of sgRNA and Cas9 gene in tomato cotyledon; however, we found that the calculated efficiency showed a large variation. It is necessary to increase the precision of the experiment to obtain reliable sgRNA efficiency data from transient expression. METHODS AND RESULTS: The cotyledon of 11^th, 15^th, 19^th, and 23^rd-day-old tomato (Solanum lycopersicum cv. Micro-Tom) were used for expressing CRISPR-Cas9 transiently. The agrobacterium harboring sgRNA for targeting ALS2 gene of tomato was injected through the stomata of leaf adaxial side and the genomic DNA was extracted in 5 days after injection. The target gene edition was identified by amplifying DNA fragment of target region and analyzing with Illumina sequencing method. The target gene editing efficiency was calculated by counting base deletion and insertion events from total target sequence read. CONCLUSION: The CRISPR-Cas9 editing efficiency varied with tomato cotyledon age. The highest efficiency was observed at the 19-day-old cotyledons. Both the median and mean were the highest at this stage and the sample variability was also minimized. We found that the transgene of CRISPR-Cas9 system was strongly correlated with plant leaf development and suggested the optimum cotyledon leaf age for Agrobacterium-mediated transfection in tomato.

• Molecules and Cells
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• 제26권5호
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• pp.474-480
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• 2008

OsMADS1 is a rice MADS box gene necessary for floral development. To identify the key cis-regulatory regions for its expression, we utilized transgenic rice plants expressing GUS fusion constructs. Histochemical analysis revealed that the 5.7-kb OsMADS1 intragenic sequences, encompassing exon 1, intron 1, and a part of exon 2, together with the 1.9-kb 5' upstream promoter region, are required for the GUS expression pattern that coincides with flower-preferential expression of OsMADS1. In contrast, the 5' upstream promoter sequence lacking this intragenic region caused ectopic expression of the reporter gene in both vegetative and reproductive tissues. Notably, incorporation of the intragenic region into the CaMV35S promoter directed the GUS expression pattern similar to that of the endogenous spatial expression of OsMADS1 in flowers. In addition, our transient gene expression assay revealed that the large first intron following the CaMV35S minimal promoter enhances flower-preferential expression of GUS. These results suggest that the OsMADS1 intragenic sequence, largely intron 1, contains a key regulatory region(s) essential for expression.

• Nuclear Engineering and Technology
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• 제52권1호
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• pp.27-36
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• 2020

Natural convection and thermal stratification phenomena are found in large water pools that are being used as heat sinks for decay heat removal from the reactor core using passive heat removal systems. In this study, the two-phase (water and air) natural convection and thermal stratification phenomena with conjugate heat transfer in the rectangular enclosure were investigated numerically using ANSYS Fluent 17.2 code. The transient numerical simulations of these phenomena in the full-scale computational domain of the experimental facility were performed. Generation of water vapour bubbles around the heater rod and evaporation phenomena were included in this numerical investigation. The results of numerical simulations are in good agreement with experimental measurements. This shows that the natural convection is formed in region above the heater rod and the water is thermally stratified in the region below the heater rod. The heat from higher region and from the heater rod is transferred to the lower region via conduction. The thermal stratification disappears and the water becomes well mixed, only after the water temperature reaches the saturation temperature and boiling starts. The developed modelling approach and obtained results provide guidelines for numerical investigations of thermal-hydraulic processes in the water pools for passive residual heat removal systems or spent nuclear fuel pools considering the concreate walls of the pool and main room above the pool.

• 한국전산유체공학회:학술대회논문집
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• 한국전산유체공학회 2001년도 추계 학술대회논문집
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• pp.99-105
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• 2001

Flow computations have been conducted to study the impingement flowfield over the KSR-III flume deflector To validate Euler solver for the jet impingement flowfieid, the jet flow over a double wedge deflector have been calculated and showed reasonable agreement with experimental data. The transient flow behavior of flume over deflector have been investigated and the flume from the rocket nozzle proved to be getting out of the deflector safely and the thermal effect on the base region of rocket was not considerable.

• 대한전자공학회:학술대회논문집
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• 대한전자공학회 1998년도 하계종합학술대회논문집
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• pp.603-606
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• 1998

In a large number of words and the continued speech recognition system using a phoneme as teh recognition unit, it is necessary to segment processing. In this paper, a normalized AMDF new method. The suggested parameter represents a degree of sharpness at valley point. This method can detect the speech segment between the steady state and transient region to the continued speech without a prior information of speech signal.

• 대한화학회지
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• 제7권1호
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• pp.47-50
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• 1963

The effect of the pulsation on the particle hold-up in a pulsed wetted column has been studied theoretically for the case of laminar region and experimentally for the cases of transient and turbulent regions. The experiment has been carried out by employing water as fluid medium and solid particles. The pulsation has been given by the reciprocating motion of a piston. The results of the investigation show that the pulsation would not give apparent effect on the particle hold-up but would increase the total travelling length of the particle.