• Title/Summary/Keyword: Complementary DNA

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Evaluation of Commercial Complementary DNA Synthesis Kits for Detecting Human Papillomavirus (인유두종바이러스 검출을 위한 상용화된 cDNA 합성 키트의 평가)

  • Yu, Kwangmin;Park, Sunyoung;Chang, Yunhee;Hwang, Dasom;Kim, Geehyuk;Kim, Jungho;Kim, Sunghyun;Kim, Eun-Joong;Lee, Dongsup
    • Korean Journal of Clinical Laboratory Science
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    • v.51 no.3
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    • pp.309-315
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    • 2019
  • Cervical cancer is the fourth most common malignant neoplasm in women worldwide. Most cases of cervical cancer are caused by an infection by the human papillomavirus. Molecular diagnostic methods have emerged to detect the HPV for sensitivity, specificity, and objectivity. In particular, real-time PCR has been introduced to acquire a more sensitive target DNA or RNA. RNA extraction and complementary DNA synthesis are proceeded before performing real-time PCR targeting RNA. To identify an adequate and sensitive cDNA synthesis kit, this study evaluated the two commonly used kits for cDNA synthesis. The results show that the $R^2$ and efficiency (%) of the two cDNA synthesis kits were similar in the cervical cancer cell lines. On the other hand, the Takara kit compared to Invitrogen kit showed P<0.001 in the $10^2$ and $10^3$ SiHa cell count. The Takara kit compared to the Invitrogen kit showed P<0.001 in the $10^1$ and $10^2$ HeLa cell count. Furthermore, 8, 4, 2, 1, and 0.5 ml of forty exfoliated cell samples were used to compare the cDNA synthesis kits. The Takara kit compared to the Invitrogen kit showed P<0.01 in 8, 4, and 1 ml and P<0.05 in 0.5 mL. The study was performed to identify the most appropriate cDNA synthesis kit and suggests that a cDNA synthesis kit could affect the real-time PCR results.

Effective Family Shuffling Method Using Complementary DNA Fragments Produced by S1 Nuclease

  • Hong, Soon-Gyu
    • Journal of Microbiology and Biotechnology
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    • v.16 no.12
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    • pp.2004-2007
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    • 2006
  • An efficient method for the in vitro reassembly of homologous DNA sequences is presented. The proposed method involves obtaining single strands of homologous genes and hybridizing them to obtain partially hybridized heteroduplex DNA; cleaving the single-stranded regions of the heteroduplex DNA using S1 nuclease to generate double-strand DNA fragments; denaturing the double-strand DNA fragments to generate single-strand DNA fragments; conducting a series of polymerase chain reactions (PCR) using the single-strand DNA fragments as internal primers and a mixture of homologous DNA as templates to obtain elongated reassembled DNA; and finally, amplifying the reassembled DNA by a PCR using terminal primers. As a result, DNA reassembly could be achieved between homologous genes with a sequence similarity as low as 78%.

Complementary DNA Cloning and Sequencing of the Coat Protein Gene of Potato Virus Y-Ordinary Korean Strain (감자바이러스 Y의 OK계통에 대한 외피단백질 유전자 cDNA 클로닝 및 염기서열 분석)

  • 정승룡;최장경;길전행이;이부영
    • Korean Journal Plant Pathology
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    • v.11 no.1
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    • pp.73-79
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    • 1995
  • Complementary DNAs (cDNAs) to the coat protein gene of an ordinary Korean strain of potato virus Y (PVY-OK) isolated from potato (cv. Superior) were synthesized and cloned into a plasmid pUC119 and sequenced. The RNA of the virus propagated in tobacco (Nicotinaa sylvestris) was extracted by the method of phenol extraction. The first strand of cDNAs to the coat protein penomic RNA of the virus was made by Moloney murine leukemia virus reverse transcriptase. The cDNA were synthesized and amplified by the method of polymerase chain reaction (PCR) using a pair of oligonucleotide primers. PVYCP3P and PVYCP3M. The size of cDNAs inserted in pUC119 plasmid was estimated as about 840 bp upon agarose gel electrophoresis. Double stranded cDNAs were transformed into the competent cell of E. coli JM109. Sequence analysis of cDNAs was conducted by the dideoxynucleotide chain termination method. Homology of cDNAs of the PVY-OK coat protein genomic RNA with those of PVY-O (Japan), PVY-T (Japan), PVY-TH (Japan), PVYN (The Netherlands),and PVYY (France) was represented as 97.3%, 88.9%, 89.3%, 89.6% and 98.5%, respectively. Homology at the amino acid level turned out to the be 97.4%, 92.5%, 92.9%, 92.9% and 98.5%, respectively.

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Development of Pattern Classifying System for cDNA-Chip Image Data Analysis

  • Kim, Dae-Wook;Park, Chang-Hyun;Sim, Kwee-Bo
    • 제어로봇시스템학회:학술대회논문집
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    • 2005.06a
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    • pp.838-841
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    • 2005
  • DNA Chip is able to show DNA-Data that includes diseases of sample to User by using complementary characters of DNA. So this paper studied Neural Network algorithm for Image data processing of DNA-chip. DNA chip outputs image data of colors and intensities of lights when some sample DNA is putted on DNA-chip, and we can classify pattern of these image data on user pc environment through artificial neural network and some of image processing algorithms. Ultimate aim is developing of pattern classifying algorithm, simulating this algorithm and so getting information of one's diseases through applying this algorithm. Namely, this paper study artificial neural network algorithm for classifying pattern of image data that is obtained from DNA-chip. And, by using histogram, gradient edge, ANN and learning algorithm, we can analyze and classifying pattern of this DNA-chip image data. so we are able to monitor, and simulating this algorithm.

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A Double Helix DNA Structure Based on Block Circulant Matrix (II) (블록순환 행렬에 의한 이중나선 DNA 구조 (II))

  • Park, Ju-Yong;Kim, Jeong-Su;Lee, Moon-Ho
    • The Journal of the Institute of Internet, Broadcasting and Communication
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    • v.16 no.5
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    • pp.229-233
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    • 2016
  • In this paper, we present the four genetic nitrogenous bases of C, U(T), A, G to matrices and describe the structures from $4{\times}4$ RNA(ribose nucleic acid) to $8{\times}8$ DNA((deoxyribose nucleic acid) matrices. we analysis a deoxyribose nucleic acid (DNA) double helix based on the block circulant Hadamard-Jacket matrix (BCHJM). The orthogonal BCHJM is anti-symmetric pair complementary of the core DNA. The block circulant ribonucleic acid (RNA) repair damage reliability is better than the conventional double helix. In case of k=4 and N=1, the reliability of block circulant complementarity is 93.75%, and in case of k=4 and N=4, it is 98.44%. Therefore it improves 4.69% than conventional case of double helix.

Highly Integrated DNA Chip Microarrays by Hydrophobic Interaction

  • Park, Yong-Sung;Kim, Do-Kyin;Kwon, Young-Soo
    • KIEE International Transactions on Electrophysics and Applications
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    • v.11C no.2
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    • pp.23-27
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    • 2001
  • Microarray-based DNA chips provide an architecture for multi-analyte sensing. In this paper, we report a new approach for DNA chip microarray fabrication. Multifunctional DNA chip microarrays were made by immobilizing many kinds if DNAs on transducers (particles). DNA chip microarrays were prepared by randomly distributing a mixture of the particles on a chip pattern containing thousands of micro meter-scale sites. The particles occupied different sites from array to array. Each particle cam be distinguished by a tag that is established on the particle. The particles were arranged on the chip pattern by the random fluidic self-assembly (RFSA) method, using hydrophobic interaction.

Structural and Dynamic Studies of the Central Segments in the Self-complementary Decamer DNA Duplexes d(ACGTATACGT)2 and d(ACGTTAACGT)2

  • Park, Jin-Young;Lee, Joon-Hwa;Choi, Byong-Seok
    • BMB Reports
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    • v.31 no.1
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    • pp.89-94
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    • 1998
  • The structures of the self-complementary decamer duplexes, $d(ACGTATACGT)_2$ (TATA-duplex) and $d(ACGTTAACGT)_2$, (TTAA-duplex) has been obtained in solution by proton NMR spectroscopy and restrained molecular dynamics. The duplexes are essentially B-type, with distortions apparent at the TATA and TTAA steps. Theses distortions and their effects on dynamics have been investigated by the measurement of imino proton exchange time of the base-pairs. The unusual opening kinetics of central A T base-pairs could be correlated to the abnormal structural properties of the corresponding sequences.

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Antiviral Efficacy of a Short PNA Targeting microRNA-122 Using Galactosylated Cationic Liposome as a Carrier for the Delivery of the PNA-DNA Hybrid to Hepatocytes

  • Kim, Hyoseon;Lee, Kwang Hyun;Kim, Kyung Bo;Park, Yong Serk;Kim, Keun-Sik;Kim, Dong-Eun
    • Bulletin of the Korean Chemical Society
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    • v.34 no.3
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    • pp.735-742
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    • 2013
  • Peptide nucleic acids (PNAs) that bind to complementary nucleic acid sequences with extraordinarily high affinity and sequence specificity can be used as antisense oligonucleotides against microRNAs, namely antagomir PNAs. However, methods for efficient cellular delivery must be developed for effective use of PNAs as therapeutic agents. Here, we demonstrate that antagomir PNAs can be delivered to hepatic cells by complementary DNA oligonucleotide and cationic liposomes containing galactosylated ceramide and a novel cationic lipid, DMKE (O,O'-dimyristyl-N-lysyl glutamate), through glycoprotein-mediated endocytosis. An antagomir PNA was designed to target miR-122, which is required for translation of the hepatitis C virus (HCV) genome in hepatocytes, and was hybridized to a DNA oligonucleotide for complexation with cationic liposome. The PNA-DNA hybrid molecules were efficiently internalized into hepatic cells by complexing with the galactosylated cationic liposome in vitro. Galactosylation of liposome significantly enhanced both lipoplex cell binding and PNA delivery to the hepatic cells. After 4-h incubation with galactosylated lipoplexes, PNAs were efficiently delivered into hepatic cells and HCV genome translation was suppressed more than 70% through sequestration of miR-122 in cytoplasm. PNAs were readily released from the PNA-DNA hybrid in the low pH environment of the endosome. The present study indicates that transfection of PNA-DNA hybrid molecules using galactosylated cationic liposomes can be used as an efficient non-viral carrier for antagomir PNAs targeted to hepatocytes.

Real-time Fluorescence Assay of DNA Polymerase Using a Graphene Oxide Platform (산화 그래핀 플랫폼을 이용한 DNA 중합효소의 실시간 형광에세이)

  • Gang, Jongback
    • Microbiology and Biotechnology Letters
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    • v.41 no.4
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    • pp.456-461
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    • 2013
  • Using the different adsorption properties of ssDNA and dsDNA to GO, this study used a real time and efficient fluorescence assay to detect the enzymatic activity of the Klenow fragment with the adsorbed DNA to GO. Results showed that adsorption of fluorescein-tagged ssDNA to GO resulted in fluorescence quenching and DNA was released from GO by adding complementary DNA. In addition, fluorescence restoration was increased through a polymerization reaction by the Klenow fragment in the presence of a fluorescein-attached template, GO, and primer. Gel electrophoresis was conducted to confirm the hybridization and DNA polymerization reactions on GO.

Detection of DNA Adduct Formed by Mitomycin C by $^{32}P$-Postlabelling ($^{32}P$-Postlabelling 방법을 이용한 미토마이신 C에 의하여 형성된 DNA adduct의 검출)

  • Jeong, Hye-Yun;Kim, Jae-Hyeon;Park, Chang-Won;Lee, Dong-Gwon
    • YAKHAK HOEJI
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    • v.40 no.4
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    • pp.442-448
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    • 1996
  • Mitomycin C(MMC) has been used as an anticancer drug and behaves as an alkylating agent forming covalent cross-link between complementary strands of double strand DNA. The purpose of this research was to determine number of DNA adducts, formed in vivo by Mitomycin C, in mouse organs. DNAs from liver, lung, brain and pancreas were isolated and used for $^{32}P$-postlabelling. The labeled nucleotides were separated by 2D-TLC and subjected to autoradiography. Numbers of MMC-DNA adducts were 9,9,5,4 in liver, pancreas, lung and brain, respectively.

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