• Title/Summary/Keyword: DNA separation

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A Molecular Phylogenetic Study on Korean Alexandrium catenella and A. tamarense Isolates (Dinophyceae) Based on the Partial LSD rDNA Sequence Data

  • Kim, Keun-Yong;Kim, Chang-Hoon
    • Journal of the korean society of oceanography
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    • v.39 no.3
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    • pp.163-171
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    • 2004
  • Sequences of the large subunit ribosomal (LSD) rDNA D1-D2 region of Alexandrium catenella(=A. sp. cf. catenella) and A. tamarense isolates, which were collected along the Korea coasts, were analyzed to understand their phylogenetic relationships and geographical distributions. All A. catenella and A. tamarense isolates belonged to the A. tamarense/catenella/fundyense complex and were grouped with the North American and temperate Asian ribotypes, respectively, regardless of the presence or absence of a ventral pore in the first apical plate. A consistent and peculiar characteristic that differentiated the Alexandrium isolates was amplification of a second PCR product with a lower molecular weight in addition to the predicted one; ten A. catenella isolates belonging to the temperate Asian ribotype yielded this additional PCR product. Sequence alignment revealed that the shorter PCR product resulted from an unusual large deletion of 87 bp in the LSD rDNA D1 domain. The North American and temperate Asian ribotypes were prevalent along the Korean coasts without geographical separation. Given the high genetic homogeneity among widely distributed Alexandrium populations, each ribotype appeared to be pandemic rather than to constitute a distinct regional population.

New Cationic Liposome with Enhanced Stability and Transfection Efficiency for Gene Delivery (안정성 및 Transfection 효율이 우수한 양이온성 리포좀 유전자 전달시스템의 개발)

  • Kim, Kyoung-Mi;Nam, Bang-Hyun;Sohn, Dong-Hwan
    • Journal of Pharmaceutical Investigation
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    • v.28 no.2
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    • pp.93-98
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    • 1998
  • We have developed liposomes which can be easily prepared with inexpensive lipid, have enhanced stability, and can efficiently deliver DNA into the COS-l cells, Liposome formulations were prepared using cationic materials such as dimethyldioctadecyl ammonium bromide (DDAB), cetyltrimethyl ammonium bromide(CTAB), We investigated the effect of cationic liposome formulations on in vitro DNA transfection, DDAB-containing liposomes showed increased transfection efficiency which was 3.2-fold as much as that by $Lipofectin^{\circledR}$, but CTAB-containing liposomes were inactive in gene transfection. The effect of colipid of DDAB-containing liposome was also investegated. As a colipid, dioleylphosphatidylethanolamine(DOPE) and cholesterol did altered the transfection efficiency of DDAB-containing liposomes. And increased DDAB concentration lowered the transfection efficiency. The optimum amount of liposomal formulation was $10\;{\mu}M$ for $1\;{\mu}g$ of DNA. In the experiment of stability, DOPE-containing liposomes formulation showed a broad size distribution and separation of two major peaks on a 5th day of preparation, but liposomes containing cholesterol was stable for 10 days. DDAB-containing liposomal DNA delivery system was prepared easily and was stable.

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Ultrastructural Studies for Protoplasts and Protoplast Fusion in Streptomyces lavendulae (Streptomyces levendulae의 원형질체와 원형질체 융합에 대한 미세구조)

  • 하영칠;홍순우;유진철;임헌만
    • Korean Journal of Microbiology
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    • v.24 no.3
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    • pp.197-203
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    • 1986
  • Morphology and ultrastructure of protoplast fusion mode in Streptomyces lavendulae were studied by scanning and transmission electron microscopy. The isolated protoplasts were stable in some degree in hypertonic solution except that several protoplasts showed irregular morphology. Fusion events were occurred as follows; contact zone, fusion zone and separation zone were appeared sequentially. After formation of the separation zone, cytoplasm and DNA from both parents were mixed eventually. In the contact zone, two menbranes were still separated by electron transparent space. The contact zone changed to fusion zone by formation of fusion membrane that phospholipid molecules of two membranes were rearranged. Thereafter, nonmembraneous separation zone was formed by disappearance of fusion membrane. These changes were characterized by successive changes in typical membrane structure in fusion areas and by a progressive loss of bispherical shape.

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12-Oxoeicosatetraenoic acid, a candidate signal for placenta separation, activates matrix metalloproteinase and induces apoptosis in bovine trophoblast cells

  • Hachiro Kamada
    • Animal Bioscience
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    • v.36 no.3
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    • pp.429-440
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    • 2023
  • Objective: 12-oxo-5Z,8Z,10E,14Z-eicosatetraenoic acid (12-KETE), a metabolite of arachidonic acid, is a strong candidate signal for placenta separation following calf discharge at delivery. In the present study, the effects of 12-KETE on bovine trophoblast cells were investigated to determine its function in the placentome at delivery. Methods: Bovine trophoblast cells derived from blastocysts were used. They were cocultured with or without fibroblasts derived from bovine placentome and/or bovine uterine epithelial cells. 12-KETE was added to the culture medium. Results: Bovine trophoblast cells contained binucleate cells and strongly expressed caudal type homeobox 2 (CDX-2) genes. Addition of 12-KETE to the trophoblast cell colony without feeder cells or that on a fibroblast monolayer induced rapid exfoliation of the colony. After 12-KETE addition, trophoblast cells emitted strong fluorescence caused by the degradation of dye-quenched collagen, indicating that 12-KETE activated matrix metalloproteinase of the trophoblast cells. Exfoliated cell colonies were stained with YOPRO-1, but not propidium iodide (PI). Moreover, DNA fragmentation and Bcl-2 associated X protein (Bax) gene (apoptosis stimulator) upregulation were observed in exfoliated cells, indicating that 12- KETE induced trophoblast cell apoptosis. These results were consistent with previous in vivo observations; however, even a lower concentration of 12-KETE activated trophoblast protease. Meanwhile, fibroblasts derived from the bovine placentome converted arachidonic acid to 12-KETE. Conclusion: These observations indicate that 12-KETE may serve as a signal for placenta separation at delivery.

Region Identification on a Trained Growing Self-Organizing Map for Sequence Separation between Different Phylogenetic Genomes

  • Reinhard, Johannes;Chan, Chon-Kit Kenneth;Halgamuge, Saman K.;Tang, Sen-Lin;Kruse, Rudolf
    • Proceedings of the Korean Society for Bioinformatics Conference
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    • 2005.09a
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    • pp.124-129
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    • 2005
  • The Growing Self-Organizing Map (GSOM), an extended type of the Self-Organizing Map, is a widely accepted tool for clustering high dimensional data. It is also suitable for the clustering of short DNA sequences of phylogenetic genomes by their oligonucleotide frequency. The GSOM presents the result of the clustering process visually on a coloured map, where the clusters can be identified by the user. This paper describes a proposal for automatic cluster detection on this map without any participation by the user. It has been applied with good success on 20 different data sets for the purpose of species separation.

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Molecular Computing Simulation of Cognitive Anagram Solving (애너그램 문제 인지적 해결과정의 분자컴퓨팅 시뮬레이션)

  • Chun, Hyo-Sun;Lee, Ji-Hoon;Ryu, Je-Hwan;Baek, Christina;Zhang, Byoung-Tak
    • KIISE Transactions on Computing Practices
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    • v.20 no.12
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    • pp.700-705
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    • 2014
  • An anagram is a form of word play to find a new word from a set of given alphabet letters. Good human anagram solvers use the strategy of bigrams. They explore a constraint satisfaction network in parallel and answers consequently pop out quickly. In this paper, we propose a molecular computational algorithm using the same process as this. We encoded letters into DNA sequences and made bigrams and then words by connecting the letter sequences. From letters and bigrams, we performed DNA hybridization, ligation, gel electrophoresis and finally, extraction and separation to extract bigrams. From the matched bigrams and words, we performed the four molecular operations again to distinguish between right and wrong results. Experimental results show that our molecular computer can identify cor rect answers and incorrect answers. Our work shows a new possibility for modeling the cognitive and parallel thinking process of a human.

Population Genetic Structure of the Korean Endemic Species, Iksookimia pacifica (Pisces: Cobitidae) Distributed in Northeast Korea (한국고유종 북방종개(어류강, 미꾸리과)의 집단유전학적 구조)

  • Jang, Sook-Jin;Ko, Myeong-Hun;Kwan, Ye-seul;Won, Yong-Jin
    • Korean Journal of Environment and Ecology
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    • v.31 no.5
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    • pp.461-471
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    • 2017
  • Population genetic studies of 10 groups of Iksookimia pacifica were conducted to investigate the genetic diversity and population genetic structure across its known range in South Korea. Population DNA sequences of one mitochondrial gene (mtCOI) and three nuclear genes (IRBP, EGR2B, RAG1) were examined in samples collected from ten streams that flow into the East Sea. Both mitochondrial and nuclear sequences exhibited significant differentiation among populations except a few cases. The Bayesian analysis of the multi-locus genotypes inferred from the DNA sequences of nuclear genes clustered the individual fish largely into two geographical groups: a northern group (from Baebong stream to Cheonjin stream) and a southern group (Yangyangnamdae stream to Gangneungnamdae stream). Given that the streams flowing into the East Sea are geographically isolated water systems, such separation of genotypes can be interpreted by the geographical separation of common ancestors into north and south that had colonized South Korea. Since the initial geographical separation of the ancestral population by north and south, the ancestral groups seem to have experienced further differentiation into the current genetic clusters through the physical isolation of streams by the East Sea in each region. It is notable that many individuals in the Jasan stream formed a genetic cluster with those of Yangyangnamdae and Gangneungnamdae streams which are distant from each other. In addition, mitochondrial gene showed low genetic differentiation between some neighboring populations and very low level of genetic diversity in several populations. The present population genetic study will provide valuable information for the conservation and management of the Korean endemic fish species, I. paicifica.

Comparison of Endonuclease-Sensitive Sites by T4 Endonuclease V and UvrABC Nuclease Treatments Followed by Formamide or Sodium Hydroxide Denaturation

  • Chang, Yung-Jin
    • BMB Reports
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    • v.31 no.4
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    • pp.405-408
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    • 1998
  • Endonuclease-sensitive sites detected by T4 endonuclease V or UvrABC nuclease treatments were compared in the dihydrofolate reductase gene of UV-irradiated Chinese hamster ovary B-11 cells. The number of endonuclease-sensitive sites detected by T4 endonuclease V treatment followed by NaOH denaturation was twice that of formamide denaturation. Repeated treatment of damaged genomic DNA with T4 endonuclease V resulted in no further increase in the number of endonuclease-sensitive sites detected. The numbers of endonuclease-sensitive sites detected by UvrABC nuclease using each denaturation condition were similar. Sequential treatment with the two endonucleases using formamide denaturation resulted in twice the number of endonuclease-sensitive sites detected by treatment of each nuclease alone. Due to a lack of AP endonuclease activity these results suggest the presence of T4 endonuclease V-sensitive sites which could be complemented by alkaline gel separation or by UvrABC nuclease treatment.

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Improved Genomic DNA Isolation from Soil (토양으로부터 genomic DNA의 효과적인 분리)

  • Kang Ju-Hyung;Kim Bo-Hye;Lee Sun-Yi;Kim Yeong-Jin;Lee Ju-Won;Park Young Min;Ahn Soon-Cheol
    • Journal of Life Science
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    • v.15 no.6 s.73
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    • pp.851-856
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    • 2005
  • Although valuable microbes have been isolated from the soil for the various productions of useful components, the microbes which can be cultivated in the laboratory are only $0.1-1\%$ of all microbes. To solve this problem, the study has recently been tried for making the valuable components from the environment by directly separating unculturable micrbial DNA in the soil. But it is known that humic acid originated from the soil interrupts various restriction enzymes and molecular biological process. Thus, in order to prevent these problems, this study modified the method separated soil DNA with phenol, CTAB and PEG. In order to compare the degree of purity for each DNA and the molecular biological application process, $A_{260}/A_{280}$ ratio, restriction enzymes, and PCR were performed. In case of DNA by the modified method, total yield of DNA was lower but $A_{260}/A_{280}$ ratio was higher than the previously reported methods. It was confirmed that the degree of purity is improved by the modified method. But it was not cut off by all kinds of tested restriction enzymes because of the operation of a very small amount of interrupting substances. When PCR was operated with each diluted DNA in different concentrations and GAPDH primer, the DNA by the modified method could be processed for PCR in the concentration of 100 times higher than by the previously reported separation method. Therefore, this experiment can find out the possibility of utilization for the unknown substances by effectively removing the harmful materials including humic acid and help establishing metagenomic DNA library from the soil DNA having the high degree of purity.

Molecular cloning, tissue distribution and quantitative analysis of two proopiomelanocortin mRNAs in Japanese flounder (Paralichthys olivaceus)

  • Kim, Kyoung-Sun;Kim, Hyun-Woo;Chen, Thomas T.;Kim, Young-Tae
    • BMB Reports
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    • v.42 no.4
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    • pp.206-211
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    • 2009
  • Proopiomelanocortin (POMC) plays an essential role in the stress response of the hypothalamic-pituitary-adrenal axis, and is the precursor of biologically active peptides such as adrenocorticotropin (ACTH), $\alpha$-melanocyte-stimulating hormone ($\alpha$-MSH), $\beta$-melanocyte-stimulation hormone ($\beta$-MSH) and $\beta$-endorphin. We have synthesized two different forms of POMC cDNA clones, POMC-I and POMC-II, from a pituitary cDNA library for Paralichthys olivaceus, or Japanese flounder. jfPOMC-I cDNA consists of 954bp and encodes a polypeptide of 216 amino acid residues, whereas jfPOMC-II consists of 971bp which encode a polypeptide of 194 amino acid residues. The high levels of jfPOMC-I and -II mRNAs detected in the pituitary tissue and moderate levels detected in the brain tissue plus our quantitative RT-PCR analysis, which showed there to be no significant difference between the levels of jfPOMC-I and -II mRNAs, indicate that there may be no functional separation between these two mRNAs in the flounder.