• Title/Summary/Keyword: Labeled DNA

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Normalization of Microarray Data: Single-labeled and Dual-labeled Arrays

  • Do, Jin Hwan;Choi, Dong-Kug
    • Molecules and Cells
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    • v.22 no.3
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    • pp.254-261
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    • 2006
  • DNA microarray is a powerful tool for high-throughput analysis of biological systems. Various computational tools have been created to facilitate the analysis of the large volume of data produced in DNA microarray experiments. Normalization is a critical step for obtaining data that are reliable and usable for subsequent analysis such as identification of differentially expressed genes and clustering. A variety of normalization methods have been proposed over the past few years, but no methods are still perfect. Various assumptions are often taken in the process of normalization. Therefore, the knowledge of underlying assumption and principle of normalization would be helpful for the correct analysis of microarray data. We present a review of normalization techniques from single-labeled platforms such as the Affymetrix GeneChip array to dual-labeled platforms like spotted array focusing on their principles and assumptions.

Turnover of Phosphate Compounds in Chlorella cells in a P-free medium (인산결핍배지에 있어서의 Chlorella 세포내의 인산화합물의 전환)

  • 이영녹
    • Journal of Plant Biology
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    • v.9 no.1_2
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    • pp.1-6
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    • 1966
  • Using the Chlorella cells which had been uniformly labeled with $^{32}P$, the distribution of phosphorus in various fractions of cell material was investigated. Uniformly $^{32}P$-labeled Chlorella cells were further grown in a P-free medium, and some protions of the cells were taken out at intervals during the culture, and subjected to analyze the contents of $^{32}P$ in various fractins of the cell constituents. 2. Analysis of the $^{32}P$-labeled Chlorella cells showed that the highest in P-content was the fraction of RNA followed by those of lipid, RNA-polyphosphate complex, acid-insoluble polyphosphate, acid-soluble polyphosphate, DNA and protein. 3. During the culture of $^{32}P$-labeled Chlorella cells in a P-free medium, amounts of phosphate in DNA, protein and lipid fractions increased, while the P-contents in the fraction of RNA-polyphosphate complex decreased as well as those of acid-insoluble polyphosphate and acid-soluble polyphosphate fractions. 4. It was inferred that phosphorus used in the syntheses of DNA and protein was taken from polyphosphates of the cells, and RNA-polyphosphate complex would play an important role as a phosphate pool.

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NMR peak assignment for the elucidation of the solution structure of T4 Endonuclease V

  • Im, Hoo-Kang;Hyungmi Lihm;Yu, Jun-Suk;Lee, Bong-Jin
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1996.04a
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    • pp.183-183
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    • 1996
  • Bacteriophage T4 endonuclease V initiates the repair of ultraviolet (UV)-induced pyrimidine dimer photoproducts in duplex DNA. The mechanism of DNA strand cleavage involves four sequential stens: linear diffusion along dsDNA, pyrimidine dimer-specific binding,l pyrimidine dimer-DNA glycosylase activity, and Af lyase activity. Although crystal structure is known for this enzyme, solution structure has not been yet known. In order to elucidate the solution structure of this enzyme NMR spectroscopy was used. As a basis for the NMR peak assignment of the protein, HSQC spectrum was obtained on the uniformly $\^$15/N-labeled T4 endonuclease V. Each amide peak of the spectrum were classified according to amino acid spin systems by interpreting the spectrum of $\^$15/N amino acid-specific labeled T4 endonuclease V. The assignment was mainly obtained from three-dimensional NMR spectra such as 3D NOESY-HMQC, 3D TOCSY-HMQC. These experiments were carried out will uniformly $\^$15/N-labeled sample. In order to assign tile resonance of backbon atom, triple-resonance theree-dimensional NMR experiments were also performed using double labeled($\^$15/N$\^$13/C) sample. 3D HNCA, HN(CO)CA, HNCO, HN(CA)HA spectra were recorded for this purpose. The results of assignments were used to interpret the interaction of this enzyme with DNA. HSQC spectrum was obtained for T4 endonuclease V with specific $\^$15/N-labeled amino acids that have been known for important residue in catalysis. By comparing the spectrum of enzyme*DNA complex with that of the enzyme, we could confirm the important role of some residues of Thr, Arg, Tyr in activity. The results of assignments were also used to predict the secondary structure by chemical shift index (CSI).

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A Versatile Method for DNA Sequencing of Unpurified PCR Products using an Automated DNA Sequencer and Tailed or Nested Primer Labeled with Near-infrared Dye: A Case Study on the Harmful Dinoflagellate Alexandrium

  • Ki Jang-Seu;Han Myung-Soo
    • Fisheries and Aquatic Sciences
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    • v.9 no.2
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    • pp.70-74
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    • 2006
  • DNA sequence-based typing is considered a robust tool for the discrimination of dinoflagellate species because of the availability of extensive rDNA sequences. Here, we present a rapid, cost-effective DNA-sequencing technique for various PCR products. This sequencing strategy relies on 'nested' or 'tailed' primer labeled with near-infrared dye, and uses a minimal volume of unpurified PCR product (ca. $5{\mu}L$) as the DNA template for sequencing reactions. Reliable and accurate base identification was obtained for several hundred PCR fragments of rRNA genes. This quick, inexpensive technique is widely applicable to sequence-based typing in clinical applications, as well as to large-scale DNA sequencing of the same genomic regions from related species for studies of molecular evolution.

Studies on the Phosphate Metabolism in Chlorella, with Special Reference to Polyphosphate (Chlorella의 인산대사에 관한 연구)

  • 이영록
    • Korean Journal of Microbiology
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    • v.2 no.1
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    • pp.1-11
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    • 1964
  • Yung Nok Lee (Dept. of Biology, Korea University) : Studies on the phosphate metabolism in Chlorella, with special reference to polyphosphate. Kor. J. Microbiol., Vol.2, No.1, p1-11 (1964). 1. Uniformly $^{32}P$-labeled Chlorella cells which were irradiated with Cobalt-60 gamma-rays of about 70, 000 $\gamma$ dose, were further grown in a standard "cold" medium ("hot".rarw."cold"), and some portions of the algae were taken out at the begining of, and at intervals during the culture, and subjected to analyze the contents of $^{32}P$- and total P in various fractions of the cell materials. Results obtained were compared with those of nonirradiated normal cells. 2. Amounts of phosphate in various fractions of the nonirradiated normal Chlorella cells were measured using uniformly $^{32}P$--labeled cells. Analysis of the $^{32}P$--labeled algal cells showed that the highest value in P-content was the fraction of RNA followed by those of lipid, polyphosphate "C" polyphosphate "B", DNA, nucleotidic labile phosphate compounds, polyphosphate "A" and protein. It was observed that content of total polyphosphates in a single Chlorella cell was almost equal to RNA-P content in the cell, and the amount of RNA-P was almost equal to ten times of DNA-P content. 3. When the $^{32}P$--labeled algae which were irradiated with gamma-rays were grown in a normal "cold" medium, phosphate contents in the fraction of DNA, nucleotidic labile phosphate compounds and protein decreased markedly, while the contents of phosphate in the fractions of polyphosphate "C" and potyphosphate "B" increased in comparison with those of unirradiated normal cells. So, it was considered that the pretreatment of above mentioned dose of gamma-ray inhibited DNA and protein synthesis from polyphosphate in Chlorella cells. 4. Proceeding the culture of $^{32}P$--labeled Chlorella in a "cold" standard medium, whose synthetic activity of DNA and protein from polyphosphate was disturded by gamma-ray irradiation, the amounts of $^{32}P$-in the fraction of polyphosphate "C" increased, in contrast with those of polyphosphate "B" fraction. According to these experimental results, it was inferred that polyphosphate "B" could transform into polyphosphate "C" in normal growing Chlorella cells.sults, it was inferred that polyphosphate "B" could transform into polyphosphate "C" in normal growing Chlorella cells.ing Chlorella cells.

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Quantification of DNA Delivery Efficiency Labeled with Fluorescent Dye in Digital Electroporation System (디지털 전기천공시스템에서 형광 염료로 표지 된 DNA 전달 효율의 정량화)

  • Bae, Seo Jun;Im, Do Jin
    • Korean Chemical Engineering Research
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    • v.58 no.3
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    • pp.450-457
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    • 2020
  • In the previous study, there was a big difference between the tendency of the delivery efficiency of Yo-Pro-1 and the expression efficiency of the CFP gene, but there was a problem that could not provide a clue to this problem. Therefore, this study aimed to present a clue to this problem by quantifying and comparing the delivery efficiency after labeling DNA using a fluorescent dye, which was one of the methods for quantifying biomolecules. As a fluorescent dye for labeling, Yo-Pro-1 was used, and the delivery efficiency of the fluorescent dye Yo-Pro-1 and the labeled DNA was compared. The delivery efficiency of Yo-Pro-1 and labeled DNA according to the voltage condition of the digital electroporation system was maximized at 96 V, and the delivery efficiency tended to decrease as the voltage increased further. In the comparison of the delivery efficiency of Yo-Pro-1 and labeled DNA according to the number of voltage application conditions, the delivery efficiency was maximized at the number of 8 voltage application times for both delivery materials, and the delivery efficiency tended to decrease as the number of voltage application increases further. Through the two results, it was confirmed that the delivery efficiency using Yo-Pro-1 in the digital electroporation system represents the delivery efficiency of the system well. In addition, through the results of this study, the difference between the tendency of the delivery efficiency of Yo-Pro-1 and the expression efficiency of the CFP gene shown in the preceding study was not the result of the difference in the delivery efficiency of the delivery material, but it can be predicted to be due to a problem with the expression process of the genetic material that had been delivered.

Detection of Japanese Encephalitis Virus by Biotinylated cDNA Probe (Biotin으로 표지된 cDNA Probe를 이용한 일본 뇌염 바이러스의 검색)

  • 황동연;신영오;임정빈
    • Korean Journal of Microbiology
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    • v.26 no.3
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    • pp.149-154
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    • 1988
  • Japanese Encephalitis Virus(JEV) can be detected conveniently by the use of biotinylated cDNA probe. To prepare biotinylated probe aminoallyl dUTP was first synthesized chemically to reverse transcribe the virial RNA. The allylamine-labeled cDNA was then converted to the biotin-cDNA by the reaction with an activated biotin ester, NHS-ACA-biotin. The JEV genomic RNA was hybridized to the biotinylated cDNA probe on nitrocellulose filter and visualized colorimetrically by streptavidin complexes with alkaline phosphatase polymer. Sensitivity of the detection system was determined by estimating the amount of the JEV genomic RNA through comparison with signals generated from the biotinylated and $^{32/P}$ -labeled probes. It was found that the biotin probe was as sensitive as $^{32/P}$ -cDNA probe which can detect 50pgs of the target RNA.

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Development of dot blot hybridization method using non-radio labeled probes for the diagnosis of malignant catarrhal fever (Dot blot hybridization에 의한 malignant catarrhal fever virus의 진단법 개발)

  • Kim, Ok-Jin
    • Korean Journal of Veterinary Pathology
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    • v.7 no.1
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    • pp.1-4
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    • 2003
  • Malignant catarrhal fever (MCF) is a systemic disease of ruminants caused by a gamma herpesvirus, ovine herpesvirus 2 (OvHV-2). Dot blot hybridization (DBH) protocols for detecting and differentiating this MCF virus were developed. OvHV-2 specific primer pairs, 556/555, were used for the amplification of target DNA. Then, the amplified DNA was labeled with incorporation of digoxigenin (DIG). The Dig-labeled probe was able to detect and differentiate specifically OvHV-2 DNA. This DBH technique can be applied to confirm the presence of MCF virus on clinical samples and to differentiate specifically between OvHV-2 infection and other viral infections.

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Radiopharmaceuticals for Imaging of Cellular Proliferation (세포 증식 영상용 방사성의약품)

  • Oh, Seung-Jun
    • The Korean Journal of Nuclear Medicine
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    • v.36 no.4
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    • pp.209-223
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    • 2002
  • By considering the biological properties of a tumor, it should be possible to realize better results in cancer therapy. PET imaging offers the opportunity to measure tumor growth non-invasively and repeatedly as an early assessment of response to cancer therapy. Measuring cellular growth instead of energy metabolism showed offer significant advantages in evaluating therapy. Thymidine and its derivative nucleoside compounds can be changed to mono, di- and tri- phosphate compounds by thymidine kinase and then be incorporated into DNA. Their bindings are increased in highly proliferating cells due to the high DNA synthesis rate. To evaluate cell proliferation, many kinds of thymidine and uridine derivatives have been labeled with positron emitter and radioactive iodine. Compared to radiopharmaceuticals which have radioisotope labeled base ring such as pyirmidine, the radiopharmacuticals which have radioisotope labeled sugar ring are more stable in vivo and have metabolic resistance. The biological properties such as DNA incorporation ratios are highly dependent on their chemical structures and metabolic processes. This overview describes synthesis of radiopharmaceuticals and their biological properties for imaging of tumor cell proliferation.

Turnover of $^{32}P$ and Total P in $^{32}P$-labeled Chlorella Cells in a Standard "Cold" Medium. ($^{32}P$-Labeled Chlorella의 정상배지에 있어서의 $^{32}P$ 및 Total P의 전환)

  • Lee, Yung-Nok;Chin, Pyung
    • Korean Journal of Microbiology
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    • v.4 no.1
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    • pp.14-20
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    • 1966
  • 1. Uniformly $^{32}P$-labeled Chlorella cells were further grown in a standard "cold" medium and aliquots of the algal cells were taken out at the beginning of, and at intervals during the culture, and subjected to analyze the contents of $^{32}$ P and total P in various fractions of the cell constituents. 2. When the $^{32}P$--labeled algae were grown in a normal "cold" medium, the P-contents in the fractions of DNA and protein increased. In the meantime the $^{32}P$- in acid-insoluble polyphosphate fraction decreased considerably, while that in RNA-polyphosphate complex significantly increased. 3. It was inferred that, under the experimental conditions of the present study, the phosphorus in polyphosphate seems to be transferred to RNA polyposphate complex and the phosphorus used in the synthesis of DNA and protein was, directly or indirectly, taken from those fractions above.ose fractions above.

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