• Title/Summary/Keyword: Enrichment Step

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Isolation and Analysis of Temperature Sensitive Mutants in Aspergillus nidulans (Aspergillus nidulans 온도감수성 돌연변이주의 분리 및 분석)

  • 박찬규;강현삼
    • Korean Journal of Microbiology
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    • v.16 no.3
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    • pp.111-121
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    • 1978
  • About 40 temperature-sensitive mutants have been isolated as a preliminary step to study the spore germination, the cell cycle, and the control of macromolecular synthesis in Aspergillus nidulans. To obtain temperature-sensitive mutants rapidly and effectively, the selective enrichment method using antifungal antibiotic nystatin was developed. Based on the data which had applied to the concentration of auxotrophic mutants by the earlier investigators, the optimal concentration and the time of treatment at the nonpermissive temeprature were determined as 50 to 100 units per ml and 4.5 hr., respectively. Out of 41 ts mutants assigned to the strain symbol PK, thirteen that seemed to be arrested at the earlystage of spore germination were subjected to the further cytological and genetic analysis. Elght of these mutants are able to form germ tube and five not. Staining with acid fuchsin for the 5PK strains shows that one irreversible mutant, PK6 strain able to form germ tube, accumulate mitotic spindle, being arrested in mitosis. Another PK15 and PK23 strain have more than one intact nucleolus without germ tube formation at the restrictive temperature. the temperature-senstive mutation in PK12 strain, the onlystrain which is able occurred in certain gene specific for the germination of spore. All of the ts markers are recessive and complement each other in heterokaryon between two different ts markers at the restrictive temperature.

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MET1-Dependent DNA Methylation Represses Light Signaling and Influences Plant Regeneration in Arabidopsis

  • Shim, Sangrea;Lee, Hong Gil;Seo, Pil Joon
    • Molecules and Cells
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    • v.44 no.10
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    • pp.746-757
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    • 2021
  • Plant somatic cells can be reprogrammed into a pluripotent cell mass, called callus, which can be subsequently used for de novo shoot regeneration through a two-step in vitro tissue culture method. MET1-dependent CG methylation has been implicated in plant regeneration in Arabidopsis, because the met1-3 mutant exhibits increased shoot regeneration compared with the wild-type. To understand the role of MET1 in de novo shoot regeneration, we compared the genome-wide DNA methylomes and transcriptomes of wildtype and met1-3 callus and leaf. The CG methylation patterns were largely unchanged during leaf-to-callus transition, suggesting that the altered regeneration phenotype of met1-3 was caused by the constitutively hypomethylated genes, independent of the tissue type. In particular, MET1-dependent CG methylation was observed at the blue light receptor genes, CRYPTOCHROME 1 (CRY1) and CRY2, which reduced their expression. Coexpression network analysis revealed that the CRY1 gene was closely linked to cytokinin signaling genes. Consistently, functional enrichment analysis of differentially expressed genes in met1-3 showed that gene ontology terms related to light and hormone signaling were overrepresented. Overall, our findings indicate that MET1-dependent repression of light and cytokinin signaling influences plant regeneration capacity and shoot identity establishment.

Distribution of Heavy Metals in Sediment Cores Collected from the Nakdong River, South Korea

  • Magalie, Ntahokaja;Lee, Jiyeong;Kang, Jihye;Kim, Jeonghoon;Park, Ho-Jin;Bae, Sang Yeol;Jeong, Seok;Kim, Young-Seog;Ryu, Jong-Sik
    • Journal of the Korean earth science society
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    • v.42 no.4
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    • pp.412-424
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    • 2021
  • Understanding the distribution of heavy metals in sediment is necessary because labile heavy metals can partition into the water column and bioaccumulate in aquatic organisms. Here we investigated six heavy metals (Co, Cu, Mn, Ni, Pb, and Zn) in sediment cores using a five-step sequential leaching method to examine the occurrence of heavy metals in the sediment. The results showed that all elements, except Mn, are depleted in the exchangeable and carbonate fractions. However, heavy metal concentrations are much higher in the Fe-Mn oxide and organic matter fractions, especially for Cu, indicating enrichment in the organic matter fraction. Furthermore, contamination parameters (contamination factor and geoaccumulation index) indicate that Mn contamination is high, primarily derived from anthropogenic sources, presenting a potential risk to ecosystems in the Nakdong River.

A Universal Analysis Pipeline for Hybrid Capture-Based Targeted Sequencing Data with Unique Molecular Indexes

  • Kim, Min-Jung;Kim, Si-Cho;Kim, Young-Joon
    • Genomics & Informatics
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    • v.16 no.4
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    • pp.29.1-29.5
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    • 2018
  • Hybrid capture-based targeted sequencing is being used increasingly for genomic variant profiling in tumor patients. Unique molecular index (UMI) technology has recently been developed and helps to increase the accuracy of variant calling by minimizing polymerase chain reaction biases and sequencing errors. However, UMI-adopted targeted sequencing data analysis is slightly different from the methods for other types of omics data, and its pipeline for variant calling is still being optimized in various study groups for their own purposes. Due to this provincial usage of tools, our group built an analysis pipeline for global application to many studies of targeted sequencing generated with different methods. First, we generated hybrid capture-based data using genomic DNA extracted from tumor tissues of colorectal cancer patients. Sequencing libraries were prepared and pooled together, and an 8-plexed capture library was processed to the enrichment step before 150-bp paired-end sequencing with Illumina HiSeq series. For the analysis, we evaluated several published tools. We focused mainly on the compatibility of the input and output of each tool. Finally, our laboratory built an analysis pipeline specialized for UMI-adopted data. Through this pipeline, we were able to estimate even on-target rates and filtered consensus reads for more accurate variant calling. These results suggest the potential of our analysis pipeline in the precise examination of the quality and efficiency of conducted experiments.

Neutronic simulation of the CEFR experiments with the nodal diffusion code system RAST-F

  • Tran, Tuan Quoc;Lee, Deokjung
    • Nuclear Engineering and Technology
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    • v.54 no.7
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    • pp.2635-2649
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    • 2022
  • CEFR is a small core-size sodium-cooled fast reactor (SFR) using high enrichment fuel with stainless-steel reflectors, which brings a significant challenge to the deterministic methodologies due to the strong spectral effect. The neutronic simulation of the start-up experiments conducted at the CEFR have been performed with a deterministic code system RAST-F, which is based on the two-step approach that couples a multi-group cross-section generation Monte-Carlo (MC) code and a multi-group nodal diffusion solver. The RAST-F results were compared against the measurement data. Moreover, the characteristic of neutron spectrum in the fuel rings, and adjacent reflectors was evaluated using different models for generation of accurate nuclear libraries. The numerical solution of RAST-F system was verified against the full core MC solution MCS at all control rods fully inserted and withdrawn states. A good agreement between RAST-F and MCS solutions was observed with less than 120 pcm discrepancies and 1.2% root-mean-square error in terms of keff and power distribution, respectively. Meanwhile, the RAST-F result agreed well with the experimental values within two-sigma of experimental uncertainty. The good agreement of these results indicating that RAST-F can be used to neutronic steady-state simulations for small core-size SFR, which was challenged to deterministic code system.

Combustion Characteristic Study of LNG Flame in an Oxygen Enriched Environment (산소부화 조건에 따른 LNG 연소특성 연구)

  • Kim, Hey-Suk;Shin, Mi-Soo;Jang, Dong-Soon;Lee, Dae-Geun
    • Journal of Korean Society of Environmental Engineers
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    • v.29 no.1
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    • pp.23-30
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    • 2007
  • The ultimate objective of this study is to develop oxygen-enriched combustion techniques applicable to the system of practical industrial boiler. To this end the combustion characteristics of lab-scale LNG combustor were investigated as a first step using the method of numerical simulation by analyzing the flame characteristics and pollutant emission behaviour as a function of oxygen enrichment level. Several useful conclusions could be drawn based on this study. First of all, the increase of oxygen enrichment level instead of air caused long and thin flame called laminar flame feature. This was in good agreement with experimental results appeared in open literature and explained by the effect of the decrease of turbulent mixing due to the decrease of absolute amount of oxidizer flow rate by the absence of the nitrogen species. Further, as expected, oxygen enrichment increased the flame temperatures to a significant level together with concentrations of $CO_2$ and $H_2O$ species because of the elimination of the heat sink and dilution effects by the presence of $N_2$ inert gas. However, the increased flame temperature with $O_2$ enriched air showed the high possibility of the generation of thermal $NO_x$ if nitrogen species were present. In order to remedy the problem caused by the oxygen-enriched combustion, the appropriate amount of recirculation $CO_2$ gas was desirable to enhance the turbulent mixing and thereby flame stability and further optimum determination of operational conditions were necessary. For example, the adjustment of burner with swirl angle of $30\sim45^{\circ}$ increased the combustion efficiency of LNG fuel and simultaneously dropped the $NO_x$ formation.

Sample Pretreatment and HRGC/HRMS Analysis for Determination of Dioxins in Environmental Samples (환경시료 중 다이옥신류의 정량을 위한 시료전처리방법의 개선 및 고분해능 GC/MS 분석에 관한 연구 Ⅰ)

  • Kim, Jin Yeong;Jang, Yun Seok;Lee, Dae Un
    • Journal of the Korean Chemical Society
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    • v.38 no.11
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    • pp.819-826
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    • 1994
  • This study described a simple, rapid and cost effective analytical technique for the ultra-trace analysis of dioxins in environmental samples. Liquid-Liquid extraction methods were used for the initial extraction and enrichment of the analyte. Subsequent clean-up procedures were achieved by using strong cation exchanger, silica and Florisil cartridges. Extracts were analysed by HRGC/HRMS-SIM. The efficiency of these analytical methods was tested by recovery and selectivity for elimination of interferences such as phenols, pesticides and PCBs in each step. The mean recovery of 1,2,3,4-TCDD spiked at 10 ppt in sea water was about 92(${\pm}$1.6)%. This analytical method was applied to Kwangyang sea water and 4.5pg/L of 2,3,7,8-TCDD was determined.

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Plant Molecular Farming Using Oleosin Partitioning Technology in Oilseeds

  • Moloney, Maurice-M.
    • Korean Journal of Plant Tissue Culture
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    • v.24 no.4
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    • pp.197-201
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    • 1997
  • Plant seed oil-bodies or oleosomes ate the repository of the neutral lipid stored in seeds. These organelles in many oilseeds may comprise half of the total cellular volume. Oleosomes are surrounded by a half-unit membrane of phospholipid into which are embedded proteins called oleosins. Oleosins are present at high density on the oil-body surface and after storage proteins comprise the most abundant proteins in oilseeds. Oleosins are specifically targeted and anchored to oil-bodies after co-translation on the ER. It has been shown that the amino-acid sequences responsible for this unique targeting reside primarily in the central hydrophobic tore of the oleosin polypeptide. In addition, a signal-like sequence is found near the junction of the hydrophobic domain and ann N-terminal hydrophilic / amphipathic domain. This "signal" which is uncleaved is also essential for correct targeting. Oil-bodies and their associated oleosins may be recovered by floatation centrifugation of aqueous seed extracts. This simple partitioning step results in a dramatic enrichment for oleosins in the oil-body fraction. In the light of these properties, we reasoned that it would be feasible to create fusion proteins on oil-bodies comprising oleosins and an additional valuable protein of pharmaceutical or industrial interest. It was further postulated that if these proteins were displayed on the outer surface of oil-bodies, it would be possible to release them from the purified oil-bodies using chemical or proteolytic cleavage. This could result in a simple means of recovering high-value protein from seeds at a significant (i.e. commercial) scale. This procedure has been successfully reduced to practice for a wide variety of proteins of therapeutic, industrial and food no. The utillity of the method will be discussed using a blood anticoagulant, hirudin, and industrial enzymes as key examples.

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Precise Synthesis of Dendron-Like Hyperbranched Polymers and Block Copolymers by an Iterative Approach Involving Living Anionic Polymerization, Coupling Reaction, and Transformation Reaction

  • Hirao Akira;Tsunoda Yuji;Matsuo Akira;Sugiyama Kenji;Watanabe Takumi
    • Macromolecular Research
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    • v.14 no.3
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    • pp.272-286
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    • 2006
  • Dendritic hyperbranched poly(methyl methacrylate)s (PMMA)s, whose branched architectures resemble the 'dendron' part(s) of dendrimer, were synthesized by an iterative methodology consisting of two reactions in each iteration process: (a) a coupling reaction of u-functionalized, living, anionic PMMA having two tert-butyldimethylsilyloxymethylphenyl(SMP) groups with benzyl bromide(BnBr)-chain-end-functionalized PMMA, and (b) a transformation reaction of the introduced SMP groups into BnBr functionalities. These two reactions, (a) and (b), were repeated three times to afford a series of dendron-like, hyperbranched (PMMA)s up to third generation. Three dendron-like, hyperbranched (PMMA)s different in branched architecture were also synthesized by the same iterative methodology using a low molecular weight, functionalized 1,1-diphenylalkyl anion prepared from sec-BuLi and 1,1-bis(3-tert-butyldime-thylsilyloxymethylphenyl)ethylene in the reaction step (b) in each iterative process. Furthermore, structurally similar, dendron-like, hyperbranched block copolymers could be successfully synthesized by the iterative methodology using $\alpha$-functionalized, living, anionic poly(2-(perfluorobutyl) ethyl methacrylate) (PRfMA) in addition to $\alpha$-functionalized, living PMMA. Accordingly, the resulting block copolymers were comprised of both PMMA and PRfMA segments with different sequential orders. After the block copolymers were cast into films and annealed, their surface structures were characterized by angle-dependent XPS and contact angle measurements. All three samples showed significant segregation and enrichment of PRfMA segments at the surfaces.

Purification and Characterization of a Protease Produced by a Planomicrobium sp. L-2 from Gut of Octopus vulgaris

  • Liu, Qing;Sun, Shujing;Piao, Meizi;Yang, Ji Young
    • Preventive Nutrition and Food Science
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    • v.18 no.4
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    • pp.273-279
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    • 2013
  • Protease widely exists in the digestive tract of animals and humans, playing a very important role in protein digestion and absorption. In this study, a high protease-producing strain Planomicrobium sp. L-2 was isolated and identified from the digestive tract of Octopus variabilis. The strain was identified by physiological and biochemical experiments and 16S rDNA sequences analysis. A protease was obtained from the strain Planomicrobium sp. L-2 through ammonium sulfate precipitation, dialysis and enrichment, DEAE-Sephadex A50 anion-exchange chromatography, and Sephadex G-100 gel chromatography. The molecular weight and properties of the protease were characterized, including optimum temperature and pH, thermal stability, protease inhibitions and metal ions. According to our results, the protease from Planomicrobium sp. L-2 strain designated as F1-1 was obtained by three-step separation and purification from crude enzyme. The molecular weight of the protease was 61.4 kDa and its optimum temperature was $40^{\circ}C$. The protease F1-1 showed a broad pH profile for casein hydrolysis between 5.0~11.0. No residual activity was observed after incubation for 40 min at $60^{\circ}C$ and 60 min at $50^{\circ}C$. F1-1 protease was inhibited by $Mn^{2+}$, $Hg^{2+}$, $Pb^{2+}$, $Zn^{2+}$, and $Cu^{2+}$ ions, as well as PMSF, indicating that the protease F1-1 was a serine protease. Additionally, research basis provided by this study could be considered for industrial application of octopus intestinal proteases.