• Clinical and Experimental Reproductive Medicine
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• v.44 no.2
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• pp.63-72
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• 2017

Objective: Hyperstimulation methods are broadly used for in vitro fertilization (IVF) in patients with infertility; however, the side effects associated with these therapies, such as ovarian hyperstimulation syndrome (OHSS), have not been well studied. N-glycoproteomes are subproteomes used for the remote sensing of ovarian stimulation in follicular growth. Glycoproteomic variation in human follicular fluid (hFF) has not been evaluated. In this study, we aimed to identify and quantify the glycoproteomes and N-glycoproteins (N-GPs) in natural and stimulated hFF using label-free nano-liquid chromatography/electrospray ionization-quad time-of-flight mass spectrometry. Methods: For profiling of the total proteome and glycoproteome, pooled protein samples from natural and stimulated hFF samples were selectively isolated using hydrazide chemistry to obtain the total proteomes and glycoproteomes. N-GPs were validated by the consensus sequence N-X-S/T (92.2% specificity for the N-glycomotif at p<0.05). All data were compared between natural versus hyperstimulated hFF samples. Results: We detected 41 and 44 N-GPs in the natural and stimulated hFF samples, respectively. Importantly, we identified 11 N-GPs with greater than two-fold upregulation in stimulated hFF samples compared to natural hFF samples. We also validated the novel N-GPs thyroxine-binding globulin, vitamin D-binding protein, and complement proteins C3 and C9. Conclusion: We identified and classified N-GPs in hFF to improve our understanding of follicular physiology in patients requiring assisted reproduction. Our results provided important insights into the prevention of hyperstimulation side effects, such as OHSS.

• Journal of Plant Biotechnology
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• v.44 no.1
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• pp.89-96
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• 2017

The CRISPR/Cas9 is a core technology that can result in a paradigm for breeding new varieties. This study describes in detail the sgRNA design, vector construction, and the development of a transgenic plant and its molecular analysis, and demonstrates how gene editing technology through the CRISPR/Cas9 system can be applied easily and accurately. CRISPR/Cas9 facilitates targeted gene editing through RNA-guided DNA cleavage, followed by cellular DNA repair mechanisms that introduce sequence changes at the site of cleavage. It also allows the generation of heritable-targeted gene mutations and corrections. Here, we present detailed procedures involved in the CRISPR/Cas9 system to acquire faster, easier and more cost-efficient gene edited transgenic rice. The protocol described here establishes the strategies and steps for the selection of targets, design of sgRNA, vector construction, and analysis of the transgenic lines. The same principles can be used to customize the versatile CRISPR/Cas9 system, for application to other plant species.

• Journal of the Korea Concrete Institute
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• v.26 no.2
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• pp.219-227
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• 2014

The purpose of this study is to investigate water purification properties of porous concrete by using effective microorganisms through the long-term continuous flow test. To solve the problems such as desorption of conventional microorganisms, in this study, tertiary treatment of the effective microorganisms identified by 16S rDNA sequence analysis was adopted per each step in the manufacturing process of porous concrete. And concentration for optimum continuous flow test and operation conditions through basic experiments according to retention time were investigated. Based on the experimental results, the porous concrete applying effective microorganisms showed no toxicity on the biological water quality and exhibited excellent removal efficiency than normal porous concrete. Therefore, contaminated water quality would be improved by treatment performance investigation of contaminants through long-term continuous flow test. If problems are complemented during the experiment process, it is expected to be able to reduce the non-point pollution sources flowing into river.

• Journal of Life Science
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• v.20 no.12
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• pp.1882-1888
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• 2010

To develop probiotics, a kind of Lactobacillus sp. was isolated from infant feces. The bacterium was identified as Lactobacillus rhamnosus through 16S rDNA sequence analysis. The strain was a facultative anaerobe which grew better in aerobic conditions. The bacterium lowered the pH of the culture solution down to 2.4 during 48 hr in the MRS medium. The strain inhibited the growth of 6 pathogens - S. aureus, L. monocytogens, S. typhimurium, E. coli O-157, V. parahaemolyticus and P. aeruginosa. When the Lactobacillus were fed to chickens, along with commercial feed, for one month, amounts of $H_2S$ and $NH_3$ in the feces of the chicken decreased to 50% and 70%, respectively, compared to those of control group chickens. Amounts of other bad smells such as $(CH_3)SH$, $(CH_3)_2S$ and $(CH_3)_2S_2$ were not much different in the Lactobacillus-fed chickens compared to the control group. On the other hand, egg weights of the chickens fed Lactobacillus were higher by about $5{\pm}1\;g$ than those in the control group.

• Journal of Microbiology and Biotechnology
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• v.12 no.4
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• pp.583-591
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• 2002

Biodegradation of fat, oil, and grease (FOGs) plays an Important role in wastewater management and water pollution control. However, many industrial food-processing and food restaurants generate FOG-containing waste waters for which there Is no acceptable technology for their pretreatment. To solve these problems, this study evaluated the feasibility of effective FOG-degrading microorganisms on the biodegradation of olive oil and FOG-containing wastewater. Twenty-two strains capable of degrading FOGs were isolated from five FOG-contaminated sites for the evaluation of their FOG degradation capabilities. Among twenty-two strains tested, the lipase-producing Pseudomonas sp. strain D2D3 was selected for actual FOG wastewater treatment. Its biodegradability was performed at 3$0^{\circ}C$ and pH 8. The extent of FOG removal efficiency was varied for each FOG tested, being the highest for olive oil and animal fat (94.5% and 94.4%), and the lowest for safflower oil (62%). The addition of organic nitrogen sources such as yeast extract, soytone, and peptone enhanced the removal efficiency of FOGs, but the addition of the inorganic nitrogen nutrients such as $NH_4$Cl and $(NH_4)_2SO_4$ did not increase. The $KH_2PO_4$ sources in 0.25% to 0.5% concentrations showed more than 90% degradability. As a result, the main pathway for the oxidation of fatty acids results in the removal of two carbon atoms as acetyl-CoA with each reaction sequence: $\beta$-oxidation. Its lipase activity showed 38.5 U/g DCW using the optimal media after 9 h. Real wastewater and FOGs were used for determining the removal efficiency by using Pseudomonas sp. strain D2D3 bioadditive. The degradation by Pseudomonas sp. strain D2D3 was 41% higher than that of the naturally occurring bacteria. This result indicated that the use of isolated Pseudomonas sp. strain D2D3 in a bioaugmentating grease trap or other processes might possibly be sufficient to acclimate biological processes for degrading FOGs.

• Parasites, Hosts and Diseases
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• v.56 no.5
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• pp.409-418
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• 2018

Acanthamoeba spp. are free-living protozoa that are opportunistic pathogens for humans. Cysteine proteases of Acanthamoeba have been partially characterized, but their biochemical and functional properties are not clearly understood yet. In this study, we isolated a gene encoding cysteine protease of A. castellanii (AcCP) and its biochemical and functional properties were analyzed. Sequence analysis of AcCP suggests that this enzyme is a typical cathepsin L family cysteine protease, which shares similar structural characteristics with other cathepsin L-like enzymes. The recombinant AcCP showed enzymatic activity in acidic conditions with an optimum at pH 4.0. The recombinant enzyme effectively hydrolyzed human proteins including hemoglobin, albumin, immunoglobuins A and G, and fibronectin at acidic pH. AcCP mainly localized in lysosomal compartment and its expression was observed in both trophozoites and cysts. AcCP was also identified in cultured medium of A. castellanii. Considering to lysosomal localization, secretion or release by trophozoites and continuous expression in trophozoites and cysts, the enzyme could be a multifunctional enzyme that plays important biological functions for nutrition, development and pathogenicity of A. castellanii. These results also imply that AcCP can be a promising target for development of chemotherapeutic drug for Acanthamoeba infections.

• Korean Journal of Microbiology
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• v.47 no.3
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• pp.231-240
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• 2011

The objective of this study was to evaluate the acid and bile tolerance, bile salt hydrolase (BSH) activity, and cholesterol assimilation ability of lactic acid bacteria isolated from mustard leaf kimchi. MLK11, MLK22, MLK27, MLK41, and MLK67 were relatively acid- and bile-tolerant strains, with more than $10^5$ CFU/ml after incubation in simulated gastric juice and intestinal fluid, while MLK53 was the most sensitive strain to acid and bile. Strains MLK22 and MLK67 deconjugated the highest level of sodium glycocholate with more than 3.5 mM of cholic acid released, while deconjugation was lowest by strains MLK13 and MLK41 which released only 1.35 mM and 1.16 mM, respectively. Specially, strains MLK22 and MLK67 showed higher deconjugation of sodium glycocholate compared to sodium taurocholate and conjugated bile mixture. Although strains MLK22 and MLK67 exhibited maximal BSH activity at the stationary phase, MLK22 had somewhat higher total BSH activity compared to MLK67 towards both sodium glycocholate and sodium taurocholate. Meanwhile, cholesterol removal varied among tested strains (p<0.05) and ranged from 5.22 to 39.16 ${\mu}g$/ml. Especially, MLK67 strain assimilated the highest level of cholesterol in media supplemented with 0.3% oxgall, cholic acid, and taurocholic acid (p<0.05). According to physiological and biological characteristics, pattern of carbohydrate fermentation, and 16S rDNA sequence, strain MLK67 that may be considered as probiotic strain due to acid and bile tolerance and cholesterol-lowering effects was identified as Pediococcus pentosaceus MLK67.

• Korean Journal of Soil Science and Fertilizer
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• v.45 no.6
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• pp.1126-1135
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• 2012

In order to produce high-quality fermenting composts, bacteria strains with high activities of extracellular enzymes (cellulase, chitinase, amylase, protease and lipase) were isolated from the soils in 6 provinces of Korea, and characterized by 16S rRNA gene sequence analysis and properties. The selected 7 stains inoculated to livestock manure for 2' fermenting time, and experimental treatment divided into 3 groups, B1, B2 and B3, according to microbial activity and enzyme type. Our results showed that microbe applications (B1, B2 and B3) can increase (p<0.05) both rhizomes (17-38%) and enzyme activities (50-81%) in compost after fermenting time, respectively, compared to non-microbe treatment (control). The microbe application also decreased significantly (p<0.05) the $NH_3$ and $H_2S$ gas contents 13.4 and 27.3% compared with control, and the Propionic acid and Butyric acid gas contents 14.5 and 19.6%, respectively, as compared to the control. The microbial degradation rate (%) of pesticides and heavy metals increased significantly (p<0.05) after fermenting time, respectively, as compared to the control. Especially, microbe applications were more effective in total rhizomes yields and bioactivities than non-microbe treatment. Thus the results of this study could help in development of potential bioinoculants and composting techniques that maybe suitable for crop production, and protectable for earth environment under various conditions.

• Horticultural Science & Technology
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• v.28 no.4
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• pp.638-647
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• 2010

The genetic relationships in 5 species of $Ligularia$ were investigated using RAPD (Randomly Amplified Polymorphic DNA) and ITS (Internal Transcribed Spacer) sequences analyses. In RAPD analysis, sixty three of 196 arbitrary primers showed polymorphism. The amplified fragments ranged from 0.2 to 1.6 kb in size. The dendrogram was constructed by the UPGMA clustering algorithm based on genetic similarity of RAPD markers. A total of 16 accessions were classified into 5 major groups corresponding each species at the similarity coefficient value of 0.77. In the ITS sequence analysis, the size of ITS 1 was varied from 248 to 256 bp, while ITS 2 was varied from 220 to 222 bp. The 5.8S coding region was 164 bp in lengths. Forty nine sites (10.2%) of the 478 nucleotides were variable, and the G+C content of ITS region ranged from 49.4 to 53.5%. In the ITS tree, five species of $Ligularia$ were monophyletic, and $L.$ $taquetii$ was the first branching within the clade. $Ligularia$ $intermedia$ formed a clade with $L.$ $fischeri$ var. $spiciformis$ (BS=79), and $L.$ $stenocephala$ and $L.$ $fischeri$ were also claded. Two data sets were congruent, except of the position of $L.$ $fischeri$ var. $spiciformis$.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.1075-1080
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• 2004

Dextransucrase (DSRB742) from Leuconostoc mesenteroides NRRL B-742CB is a glucosyltransferase that catalyzes the synthesis of dextran using sucrose, or the synthesis of oligosaccharides when acceptor molecules, like maltose, are present. The DSRB742 gene (dsrB742) was cloned and the properties were characterized. In order to identify critical amino acid residues, the DSRB742 amino acid sequence was aligned with glucosyltransferase sequences, and three amino acid residues reported as sucrose binding amino acids in Streptococcus glucosyltransferases were selected for site-directed mutagenesis experiments. Asp-533, Asp-536, and His-643 were independently replaced with Ala or Asn. D533A and D536A dextransucrases showed reduced dextran synthesis activities, 2.3% and 40.8% of DSRB742 dextransucrase, respectively, and D533N, D536N, H643A, end H643N dextransucrases showed complete suppression of dextran synthesis activities altogether. Additionally, D536N dextransucrase showed complete suppression of oligosaccharide synthesis activities. However, modifications at Asp-533 or at His-643 retained acceptor reaction activities in the range of 8.4% to 21.3% of DSRB742 acceptor reaction activity. Thus at least two carboxyl groups of Asp-533 and Asp-536, and His-643 as a proton donor, are essential for the catalysis process.