• Title/Summary/Keyword: Molecular Detection

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Immunochemical Studies of Starfish Gangliosides: Production of Monoclonal Antibody against AG-2, the Major Ganglioside of Starfish Acanthaster planci, and Detecting Its Distribution in Tissues by TLC Immunostaining

  • Miyamoto, Tomofumi;Yamamoto, Atsushi;Sakai, Maki;Tanaka, Hiroyuki;Shoyama, Yukihiro;Higuchi, Ryuichi
    • Journal of Marine Bioscience and Biotechnology
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    • v.1 no.4
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    • pp.298-304
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    • 2006
  • In this study, we establish a thin-layer chromatography (TLC) immunostaining method for detecting starfish gangliosides. A new monoclonal antibody (MAb) against AG-2, the major gangliosides molecular species of Acanthaster planci, was produced by fusing hybridoma with splenocytes immunized to liposomal AG-2. BALB/c male mice were injected with liposomal AG-2 antigen, and immunized. Their splenocytos were isolated and fused with hypoxanthine-aminopterine-thimidine (HAT)-sensitive mouse myeloma cells. Hybridomas producing MAb reactive to AG-2 were cloned using the limited dilution method. Established hybridomas were cultured in eRDF medium. Crude MAb produced from clone 8D4 was purified with a magnesium pyrophosphate column. Enzyme immunoassay and TLC immunostaining of AG-2 were performed using the purified MAb. Structurally related gangliosides did not cross-react with anti-AG-2 antibodies. The detection limit of TLC immunostaining was 50 ng of AG-2. The newly established immunostaining method was further developed for detecting AG-2 distribution and qualitative analysis in tissues and/or organs. Our results show that the majority of AG-2 is present in the stomach of male A. planci, while AG-2 is distributed not only in the stomach but also in the the pyloric caeca of female A. planci.

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Improved Resistance to Oxidative Stress by a Loss-of-Function Mutation in the Arabidopsis UGT71C1 Gene

  • Lim, Chae Eun;Choi, Jung Nam;Kim, In A;Lee, Shin Ae;Hwang, Yong-Sic;Lee, Choong Hwan;Lim, Jun
    • Molecules and Cells
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    • v.25 no.3
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    • pp.368-375
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    • 2008
  • Approximately 120 UDP-glycosyltransferases (UGTs), which are classified into 14 distinct groups (A to N), have been annotated in the Arabidopsis genome. UGTs catalyze the transfer of sugars to various acceptor molecules including flavonoids. Previously, UGT71C1 was shown to glycosylate the 3-OH of hydroxycinnamates and flavonoids in vitro. Such secondary metabolites are known to play important roles in plant growth and development. To help define the role of UGT71C1 in planta, we investigated its expression patterns, and isolated and characterized a loss-of-function mutation in the UGT71C1 gene (named ugt71c1-1). Our analyses by quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR), microarray data mining, and histochemical detection of GUS activity driven by the UGT71C1 promoter region, revealed the tissue-specific expression patterns of UGT71C1 with highest expression in roots. Interestingly, upon treatment with methyl viologen (MV, paraquat), ugt71c1-1 plants displayed enhanced resistance to oxidative stress, and ROS scavenging activity was higher than normal. Metabolite profiling revealed that the levels of two major glycosides of quercetin and kaempferol were reduced in ugt71c1-1 plants. In addition, when exposed to MV-induced oxidative stress, eight representative ROS response genes were expressed at lower levels in ugt71c1-1 plants, indicating that ugt71c1-1 probably has higher non-enzymatic antioxidant activity. Taken together, our results indicate that ugt71c1-1 has increased resistance to oxidative stress, suggesting that UGT71C1 plays a role in some glycosylation pathways affecting secondary metabolites such as flavonoids in response to oxidative stress.

Effects of Au Nanoparticle Monolayer on or Under Graphene for Surface Enhanced Raman Scattering

  • Kim, B.Y.;Jung, J.H.;Sohn, I.Y.;Lee, N.E.
    • Proceedings of the Korean Vacuum Society Conference
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    • 2013.02a
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    • pp.636-636
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    • 2013
  • Since first discovery of strong Raman spectrum of molecules adsorbed on rough noble metal, surface enhanced Raman scattering (SERS) has been widely used for detection of molecules with low concentration. Surface plasmons at noble metal can enhance Raman spectrum and using Au nanostructures as substrates of SERS has advantages due to it has chemical stability and biocompatibility. However, the photoluminescence (PL) background from Au remains a problem because of obtaining molecular vibration information. Recently, graphene, two-dimensional atomic layer of carbon atoms, is also well known as PL quenchers for electronic and vibrational excitation. In this study, we observed SERS of single layer graphene on or under monolayer of Au nanoparticles (NPs). Single layer graphene is grown by chemical vapor deposition and transferred onto or under the monolayer of Au NPs by using PMMA transfer method. Monolayer of Au NPs prepared using Langmuir-Blodgett method on or under graphene surface provides closed and well-packed monolayer of Au NPs. Scanning electron microscopy (SEM) and Raman spectroscopy (WItec, 532 nm) were performed in order to confirm effects of Au NPs on enhanced Raman spectrum. Highly enhanced Raman signal of graphene by Au NPs were observed due to many hot-spots at gap of closed well-packed Au NPs. The results showed that single layer graphene provides larger SERS effects compared to multilayer graphene and the enhancement of the G band was larger than that of 2D band. Moreover, we confirm the appearance of D band in this study that is not clear in normal Raman spectrum. In our study, D band appearance is ascribed to the SERS effect resulted from defects induced graphene on Au NPs. Monolayer film of Au NPs under the graphene provided more highly enhanced graphene Raman signal compared to that on the graphene. The Au NPs-graphene SERS substrate can be possibly applied to biochemical sensing applications requiring highly sensitive and selective assays.

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Anticancer Activity of Sulfated Polysaccharides Isolated from the Antarctic Red Seaweed Iridaea cordata

  • Kim, Hak Jun;Kim, Woo Jung;Koo, Bon-Won;Kim, Dong-Woo;Lee, Jun Hyuck;Nugroho, Wahyu Sri Kunto
    • Ocean and Polar Research
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    • v.38 no.2
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    • pp.129-137
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    • 2016
  • This study aimed to isolate and characterize sulfated polysaccharides (SPs) from Iridaea cordata and evaluate their anticancer activity. SPs of the Antarctic red seaweed were obtained by $CaCl_2$ (SP1) and ethanol precipitations (SP2) following diluted acid extraction at room temperature. Yields of SP1 and SP2 were approximately 14% and 23%, respectively, of the dry weight of red seaweed. The average molecular mass of the SP1 and SP2 was estimated about $1.84{\times}10^3$ and $1.42{\times}10^3kDa$, respectively, by size-fractionation High-Performance Liquid Chromatography (HPLC). From the High-Performance Anion-Exchange Chromatography-Pulsed Amperometric Detection (HPAEC-PAD) analysis, the main monosaccharide was galactose with glucose and fucose as minor components. The sulfate content of SP2 (40.4%) was slightly higher than that of SP1 (33.8%). The FT-IR spectra also showed characteristic band of carrageenan-like sulfated polysaccharides. Taken together the SPs are thought to be carrageenan-like sulfated galactan. The polysaccharides (SPs) from I. cordata exhibited weak antitumor activity against PC-3 (prostate cancer), HeLa (cervical cancer), and HT-29 (human colon adenocarcinoma). To our knowledge, this is the first data on biological activity of the Antarctic red seaweed I. cordata.

Spectral Bio-signature Simulation of full 3-D Earth with Multi-layer Atmospheric Model and Sea Ice Coverage Variation

  • Ryu, Dong-Ok;Seong, Se-Hyun;Lee, Jae-Min;Hong, Jin-Suk;Jeong, Soo-Min;Jeong, Yu-Kyeong;Kim, Sug-Whan
    • Bulletin of the Korean Space Science Society
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    • 2009.10a
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    • pp.48.1-48.1
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    • 2009
  • In recent years, many candidates for extra-solar planet have been discovered from various measurement techniques. Fueled by such discoveries, new space missions for direct detection of earth-like planets have been proposed and actively studied. TPF instrument is a fair example of such scientific endeavors. One of the many technical problems that space missions such as TPF would need to solve is deconvolution of the collapsed (i.e. spatially and temporally) spectral signal arriving at the detector surface and the deconvolution computation may fall into a local minimum solution, instead of the global minimum solution, in the optimization process, yielding mis-interpretation of the spectral signal from the potential earth-like planets. To this extend, observational and theoretical understanding on the spectral bio-signal from the Earth serves as the key reference datum for the accurate interpretation of the planetary bio-signatures from other star systems. In this study, we present ray tracing computational model for the on-going simulation study on the Earth bio-signatures. A multi-layered atmospheric model and sea ice variation model were added to the existing target Earth model and a hypothetical space instrument (called AmonRa) observed the spectral bio-signals of the model Earth from the L1 halo orbit. The resulting spectrums of the Earth show well known "red-edge" spectrums as well as key molecular absorption lines important to harbor life forms. The model details, computational process and the resulting bio-signatures are presented together with implications to the future study direction.

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Molecular Characterization of Plasmids Encoding CTX-M β-Lactamases and their Associated Addiction Systems Circulating Among Escherichia coli from Retail Chickens, Chicken Farms, and Slaughterhouses in Korea

  • Jo, Su-Jin;Woo, Gun-Jo
    • Journal of Microbiology and Biotechnology
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    • v.26 no.2
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    • pp.270-276
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    • 2016
  • Extended-spectrum β-lactamases (ESBLs), particularly those of the CTX-M types, are the predominant resistance determinants of Escherichia coli that are rapidly spreading worldwide. To determine CTX-M types, E. coli isolates were collected from retail chickens (n = 390) and environmental samples from chicken farms (n = 32) and slaughterhouses (n = 67) in Korea. Fifteen strains harboring blaCTX-M genes were isolated from 358 E. coli isolates. The most common CTX-M type was eight of CTX-M-15, followed by six of CTX-M-1 and one of CTX-M-14. The blaCTX-M genes were identified in the isolates from retail chickens (n = 9), followed by feces, water pipes, floors, and walls. Conjugations confirmed the transferability of the plasmids carrying blaCTX-M genes to the recipient E. coli J53 strain. Furthermore, eight addiction systems carried by the replicons in CTX-M types were confirmed. The dominant system was identified as ccdAB, vagCD, and pndAC in donor strains and transconjugants. The clonal relationship between the two strains carrying blaCTX-M genes indicates that E. coli may transmit from the farm to retail chickens, suggesting a possible public health risk. Our findings demonstrate that the detection of CTX-M types in E. coli isolates is important for tracking ESBL production in animals, and suggest linkage of multiple addiction systems in plasmids bearing blaCTX-M genes.

Biodegradation of Di-n-Butyl Phthalate by Rhodococcus sp. JDC-11 and Molecular Detection of 3,4-Phthalate Dioxygenase Gene

  • Jin, De-Cai;Liang, Ren-Xing;Dai, Qin-Yun;Zhang, Rui-Yong;Wu, Xue-Ling;Chao, Wei-Liang
    • Journal of Microbiology and Biotechnology
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    • v.20 no.10
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    • pp.1440-1445
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    • 2010
  • Rhodococcus sp. JDC-11, capable of utilizing di-n-butyl phthalate (DBP) as the sole source of carbon and energy, was isolated from sewage sludge and confirmed mainly based on 16S rRNA gene sequence analysis. The optimum pH, temperature, and agitation rate for DBP degradation by Rhodococcus sp. JDC-11 were 8.0, $30^{\circ}C$, and 175 rpm, respectively. In addition, low concentrations of glucose were found to inhibit the degradation of DBP, whereas high concentrations of glucose increased its degradation. Meanwhile, a substrate utilization test showed that JDC-11 was also able to utilize other phthalates. The major metabolites of DBP degradation were identified as monobutyl phthalate and phthalic acid by gas chromatography-mass spectrometry, allowing speculation on the tentative metabolic pathway of DBP degradation by Rhodococcus sp. JDC-11. Using a set of new degenerate primers, a partial sequence of the 3,4-phthalate dioxygenase gene was obtained from JDC-11. Moreover, a sequence analysis revealed that the phthalate dioxygenase gene of JDC-11 was highly homologous to the large subunit of the phthalate dioxygenase from Rhodococcus coprophilus strain G9.

Occurrence and Molecular Characterization of Noroviruses in Korean Surface Water Between 2007 and 2010

  • Lee, Gyu-Cheol;Kim, Min-Jeong;Kim, Jong Ik;Lee, Chan Hee
    • Journal of Microbiology and Biotechnology
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    • v.24 no.4
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    • pp.556-562
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    • 2014
  • The occurrence of human norovirus (NoV) genogroup I (GI) and genogroup II (GII) strains was investigated in Korea. Between 2007 and 2010, 265 samples were collected from 89 Korean water source locations. NoV GI was detected in 4.5% and NoV GII in 1.5%. Samples collected in winter had the highest occurrence; 9.4% for NoV GI and 6.3% for NoV GII. NoV GI detection was highest in groundwater, with the next highest in river water and the lowest in lake water (5.9%, 5.4%, and 1.6%, respectively), and NoV GII was found only in river water. When three representative Korean basin systems (Han (H)-, Geum/Seom (G/S)-, and Nakdong (N)-river basins) were compared, both NoV genogroups were high in the G/S-, but absent in the H- river basin. The most prevalent genotypes within the GI and GII groups were GI.5 and GII.4, respectively. The NoVs found in surface water were identical to those found in patients and those found in groundwater. The NoVs appeared to be transmitted from the patient to the surface water, and then to the groundwater, suggesting a fecal-oral route of transmission. This is the first nationwide surveillance of NoV in major Korean water sources.

The Evaluation of the thermal degradation and the degree of cure of glass/epoxy composite by ultrasonic technique (복합재료의 열화도 및 경화도에 따른 초음파 특성 연구)

  • 강길호;최원종;박상윤
    • Composites Research
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    • v.16 no.6
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    • pp.33-40
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    • 2003
  • The initial thermal degradation of polymer matrix composite is not observed easily. At the beginning of thermal degradation of polymer matrix composites, phase transformation such as chain scission, oxidation occur, and then micro delamination is produced in matrix and interface between matrix and fiber before blistering. Initial heat damage deteriorate mechanical properties of composites. We presented the detection method of the initial heat damage of composites conveniently using ultrasonic technique. Absorption coefficient and material velocity was measured with thermal degradation and degree of cure. The more thermal degradation was progressed, the more absorption coefficient was increased. When the cure temperature is more high, the absorption coefficient of cured composite is increased and material velocity is decreased. We concluded that cure temperature is more high, the defects such as void is increased and molecular structure cured at high temperature has cross-linking structure which is more absorb the ultrasonic waves.

Molecular Identification of Korean Mountain Ginseng Using an Amplification Refractory Mutation System (ARMS)

  • In, Jun-Gyo;Kim, Min-Kyeoung;Lee, Ok-Ran;Kim, Yu-Jin;Lee, Beom-Soo;Kim, Se-Young;Kwon, Woo-Seang;Yang, Deok-Chun
    • Journal of Ginseng Research
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    • v.34 no.1
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    • pp.41-46
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    • 2010
  • Expensive herbs such as ginseng are always a possible target for fraudulent labeling. New mountain ginseng strains have occasionally been found deep within mountain areas and commercially traded at exorbitant prices. However, until now, no scientific basis has existed to distinguish such ginseng from commonly cultivated ginseng species other than by virtue of being found within deep mountain areas. Polymerase chain reaction (PCR) analysis of the internal transcribed spacer has been shown to be an appropriate method for the identification of the most popular species (Panax ginseng) in the Panax ginseng genus. A single nucleotide polymorphism (SNP) has been identified between three newly found mountain ginseng (KGD4, KGD5, and KW1) and already established Panax species. Specific PCR primers were designed from this SNP site within the sequence data and used to detect the mountain ginseng strains via multiplex PCR. The established multiplex-PCR method for the simultaneous detection of newly found mountain ginseng strains, Korean ginseng, and foreign ginseng in a single reaction was determined to be effective. This study is the first report of scientific discrimination of "mountain ginsengs" and describes an effective method of identification for fraud prevention and for uncovering the possible presence of other, cheaper ginseng species on the market.