• Title/Summary/Keyword: DNA separation

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High Throughput Magnetic Separation for Human DNA by Aminosilanized Iron Oxide Nanoparticles (아미노실란화 철산화물 나노입자를 이용한 Human DNA의 초고속 자성분리)

  • Kang, Ki-Ho;Chang, Jeong-Ho
    • Journal of the Korean Ceramic Society
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    • v.45 no.10
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    • pp.605-609
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    • 2008
  • This work describes the preparation of functionalized magnetic nanoparticles(MNPs) and their bioapplication to human DNA separation. Silica coated MNPs were prepared by changing the volume ratio of tetraethyl orthosilicate(TEOS) for controlled coating thickness on the original nanoparticle of MNPs. The sol-gel process in silica coating on MNPs surface was adapted for relatively mild reaction condition, low-cost, and surfactant-free. And then amino functionalized magnetic nanoparticles were synthesized using amine groups as surface modifiers. The result of adsorption efficiency for human DNA with amino-functionalized silica coated MNPs was calculated as a function of the number of amine groups.

Capillary Electrophoresis of Single-stranded DNA

  • Choi, Hyun-Ju;Kim, Yong-Seong
    • Bulletin of the Korean Chemical Society
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    • v.24 no.7
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    • pp.943-947
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    • 2003
  • We have studied the migration behavior of single-stranded DNA using capillary gel electrophoresis under various conditions. It was found that optimum electric fields should be less than 150 V/cm for the good tradeoff between the separation time and the resolution. It seems that the gel matrix with the combination of different polymer average molecular weights is important to extend the maximum readable DNA bases. The total gel concentration less than 3.1% in the mixed gel system showed good separation efficiency up to 600 bases. The best result was obtained with the poy(ethylene)oxide (PEO) gel concentration of 1.2% of Mr 8,000,000 and 1.8% of Mr 600,000. We observed that the capillary length between 50 cm to 100 cm (effective length) should be employed for the optimization between the total DNA migration time and the maximum readable length. A trizma base-boric acid-ethlyenediaminetetraacetic acid (EDTA) (TBE) buffer was commonly used for DNA sequencing, but we found that 3-[tris(hydroxymethyl)methyl amino]-1-propane sulfonic acid (TAPS) buffer worked as well for the single-stranded DNA separation. Especially, TAPS buffer showed a good resolution for very short DNA bases (1 to 30 bases).

A DNA Microextractor Using Crossed Field Electrophoresis (교차 전기영동법을 이용한 극소형 DNA 추출기)

  • Yi Soyeon;Seo Kyoung-Sun;Cho Young-Ho
    • Transactions of the Korean Society of Mechanical Engineers A
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    • v.28 no.8 s.227
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    • pp.1135-1139
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    • 2004
  • This paper presents a microextractor for the separation of DNA molecules by their sizes. The DNA microextractor immobilizes the DNA molecules of specific size in the micropillar array by adjusting the period of the crossed electric field, thus providing a starting-point independent target DNA extraction method without separation process monitoring. The DNA microextractor has been fabricated by a three-mask micromachining process. The velocity of three different DNA molecules has been measured at the electric field of E=5V/0.8cm in the fabricated DNA microextractor, resulting in the reorientation times of $4.80{\pm}0.44sec,\;7.12{\pm}0.75sec$, and $9.88{\pm}0.30sec$ for ${\lambda}$ DNA, micrococcus DNA, and T4 DNA, respectively. T4 DNA is trapped in the micropillar array when the crossed electric field of 5V/0.8cm is applied alternately at a 10 second time interval. The present DNA microextractor filters the DNA in a specific size range by adjusting the magnitude and/or the period of the crossed electric field applied in the micropillar array.

DNA Separation Using Cellulose Derivatives and PEO by PDMS Microchip

  • Kang, Chung-mu;Back, Seung-Kwon;Song, In-gul;Choi, Byung-ok;Chang, Jun-keun;Cho, Keun-chang;Kim, Yong-seong
    • Bulletin of the Korean Chemical Society
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    • v.27 no.4
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    • pp.519-523
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    • 2006
  • Poly(dimethyl siloxane) (PDMS) has been employed as a microchip material for DNA separation in microfluidic condition. Different sieving molecules such as cellulose derivatives having glucose building block (methyl cellulose (MC), hydroxyethyl cellulose (HEC), and hydroxypropyl methyl cellulose (HPMC)) and polyethylene oxide (PEO) having linear (ring-opened ethylene oxide) unit were used and their performance was compared in terms of separation efficiency and resolution. In general, PEO showed better separation performance than cellulose derivatives probably due to the nature of linear shape polymer conformation. It was possible to perform at least 15 consecutive running with 1.2% PEO at the electric field strength around 200 V/cm. Fast analysis of the standard $\Phi$X 174 RF DNA/Hae III (less than 130s) was obtained with the number of the theoretical plate around 250,000/m. Our PMDS microchip was applied to the measurement of CAG repeat number, which is related to male infertile disease.

Development of a Method for Rapid Analysis of DNA Hybridization (측방유동방식 신속 DNA 교잡 분석법의 개발)

  • 정동석;최의열
    • Korean Journal of Microbiology
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    • v.39 no.2
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    • pp.114-117
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    • 2003
  • In molecular biology, it is necessary to develop an easy and rapid method to identify a specific DNA sequence. Though Southern and Northern blot techniques have been used widely for the analysis of gene structure and function, those methods are inconvenient in the points that we need to control incubation temperature, time, and other parameters to get the final result. In this study, we report a new method for the rapid analysis of specific DNA sequence with the modification of an immunochromatographic method. The lateral flow DNA analysis strip is composed of a sample pad, a nitrocellulose membrane for the separation and propagation of analytes, and an absorption pad for the generation of capillary action. Capture DNA was immobilized on the membrane by UV cross-linking and target DNA was labeled with Cy-5 for signaling. The samples containing target DNA were applied onto the sample pad, incubated for 15 min for separation, and scanned with a GSI fluorescence scanner. Though the hybridization reaction occurs in a short time without any washing steps, there appears to be little cross hybridization between the different sequences. The result showed a possibility that the new method can be used for the rapid identification of specific DNA sequence among the samples.

Selective DNA Adsorption on Layered Double Hydroxide Nanoparticles

  • Kim, Kyoung-Min;Park, Chung-Berm;Choi, Ae-Jin;Choy, Jin-Ho;Oh, Jae-Min
    • Bulletin of the Korean Chemical Society
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    • v.32 no.7
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    • pp.2217-2221
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    • 2011
  • We investigated the selective deoxyribonucleic acid (DNA) adsorption on layered double hydroxide (LDH) nanoparticles via studying the interaction between positively charged LDH nanoparticle as adsorbent and negatively charged adsorbates such as methyl orange (MO), fluorescein (FL), and DNA strands. The size controlled LDH $(Mg_{0.78}Al_{0.22}(OH)_2(CO_3)_{0.11}{\cdot}mH_2O)$ was prepared by conventional coprecipitation method, followed by the hydrothermal treatment. According to the adsorption isotherms, the adsorbed amounts of MO and FL were similar, however, that of DNA were much larger. The adsorption behaviors were well fitted to Freundlich adsorption model. The concentration dependent adsorption behavior on LDH surface was described in order to verify the selective DNA separation ability. The result showed that the LDH has advantages in selective adsorption of DNA competing with single molecular anions.

Separation Study of Cytosine and Guanine by HPLC and Aspen Chromatography (Aspen Chromatography 전산모사와 HPLC를 이용한 구아닌 시토신의 분리특성연구)

  • Park, Moon Bae;Kim, In Ho
    • Korean Chemical Engineering Research
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    • v.48 no.1
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    • pp.88-92
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    • 2010
  • DNA structure studies attract many interests in pharmaceutical, biochemical and medical disciplines. Among them, base pairs play a vital role in biological information transfer. Therefore, they need to be analyzed in various ways and the pair of guaninine and cytosine is the present analytical object. Separation of guanine and cytosine was researched by Aspen chromatography simulator and HPLC(High Performance Liquid Chromatography) experiments. Aspen chromatography simulation resulted in various chromatograms with changes of sample concentration, eluent flow rate and number of plate. The resolutions and yields of guanine and cytosine were calculated to obtain a best separation condition. $C_{18}$ HPLC column and water/methanol/acetic acid mixture(90/10/0.2) were used for separation of guanine and cytosine. HPLC parameters(resolution and number of theoretical plate) were calculated under different flow rates and sample concentrations. Aspen chromatography simulation and HPLC experimental results were compared with fair agreement.

High-Performance liquid Chrmatogrphic and Tandem Mass Spectrometric Quantitation of N7-Methyldeoxyguanosin in Methylated Calf Thymus DNA

  • Chae, Whi-Gun
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.5 no.3
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    • pp.191-195
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    • 2000
  • Quantitation of N7-methyldeoxyguanosine (N7-MedG) produced in the in vitro N-methly-N-nitrosuourea (NMU) action on thymus DNA has been achieved by enzymatic degradation, liquid chromatoraphic separaphic separation and desorption chemical ionization tandem mass spectrometry. In conjunction with the resolving power HPLC in the separation of isomers, desorprion chemical ionization tandem mass spectrometry has utilized in determining modified nucleosides at low levels using a stable-isotope labled compound as an internal reforence. The quantitative estimation of N7-methyldeoxyguanosine was previously established by an independent HPLC analysis of methylated calf thymus DNA. A sensitive and specific methodogy for the quantitation of N7-MedG at the picomole level using HPLC combined with tandem mass spectrometry without radioisotope labeling process is presented. The potential of the liquid chromatoraphic tandem mass exposure to methlation agents in vitro.

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Fluorescence Microscopy of Condensed DNA Conformations of Bacterial Cells

  • Suleymanoglu, Erhan
    • Journal of Microbiology
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    • v.40 no.4
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    • pp.319-326
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    • 2002
  • Cellular DNA in prokaryotes is organized in nucleic acid-protein self-assemblies referred to as the nucleoid. The physical forces responsible for its stability inside the poor solvent properties of the cytoplasm and their functional implications are not understood. Studies on the organisation and functioning of the cytosol of cells largely rely on experimental protocols performed in highly dilute solutions using biochemically purified molecules, which is not a reliable substitute for the situation existing in vivo. Our current research interest is focused on the characterization of biological and physical forces determining the compaction and phase separation of DNA in Escherichia coli cytoplasm. We have emphasized the effect of excluded volume in solutions with high macromolecular concentrations (macromolecular crowding) upon self-association patterns of reactions. The prokaryotic cytosol was simulated by addition of inert polymer polyethylene glycol (PEG) (average molecular weight 20000), as an agent which afterwards facilitates the self-association of macromolecules. Fluorescence microscopy was used for direct visualization of nucleoids in intact cells, after staining with DAPI (4',6-diamidino-2-phenylindole dihydrochloride). Addition of the crowding agent PEG 20,000, in increasing concentrations generated progressively enhanced nucleoid compaction, the effect being stronger in the presence of 0.2 M NaCl and 5 mM MgCl$\_$2/. Under these conditions, the nucleoids were compacted to volumes of around 2 ㎛$\^$3/ or comparable sizes with that of living cells.

DNA Barcoding of Boccardiella hamata (Annelida: Polychaeta: Spionidae) in South Korea

  • Lee, Geon Hyeok;Yoon, Seong Myeong;Min, Gi-Sik
    • Animal Systematics, Evolution and Diversity
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    • v.36 no.3
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    • pp.268-273
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    • 2020
  • A spionid polychaete, Boccardiella hamata (Webster, 1879) has been found from mud in crevices between the shells of oysters and adherent substrates in South Korea. The sequences of mitochondrial DNA (mtDNA) cytochrome c oxidase subunit 1 (CO1), 16S ribosomal DNA (16S), and the nuclear 18S ribosomal DNA (18S) from Korean individuals of Boccardiella hamata were determined in the present study. The molecular analysis based on the 18S rRNA gene sequences showed clear separation among the spionid polychaete species, and the sequences of Korean and Japanese individuals are completely identical. The morphological diagnosis and photographs of B. hamata are also provided.