• Title/Summary/Keyword: Cloning

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Molecular Cloning of the Gene Coding for 3-Isopropylmalate Dehydrogenase of Kluyveromyces fragilis (Kluyveromyces fragilis의 LEU gene의 Cloning)

  • 박성희;이동선;우주형;김종국;홍순덕
    • Microbiology and Biotechnology Letters
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    • v.18 no.3
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    • pp.305-308
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    • 1990
  • In order to clone the gene coding for 3-isopropylmalate dehydrogenase of Muyveromyces fragilis, a shuttle plasmid vector pHNll4 was used. It can serve as a cloning vector in Saccharomyces cerevisiae DBY746 for other Sau3AI-cleaved DNA segment of Kluyveromyces fragilis. Two cloned fragments which complement the leu2 mutation of Saccharomyces cerevisiae and E, coli were obtained. Their length was 4.4 kb an 3.5 kb, and their orientation was opposite each other. From the fact that the two recombinant plasmids were expressed in Saccharomyces cerevisiae and E, coli, probably the two inserts had the promoter of Ktuyveromyces fi-agilis and that of Kluyveromyces fiagilis was efficiently assosiated with RNA polymerase of Saccharomyces cerevisiae and E. coli. According to the result of Southern hybridization, we thought that the cloned fragment has low homology with 3-isopropylmalate dehydrogenase coding region of E. coli and Saccharomyces cerevisiae.

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Positional Cloning and Phenotypic Characterization of a New Mutant Mouse with Neuronal Migration Abnormality

  • Park, Chankyu;Ackerman, Susan-L
    • Proceedings of the KSAR Conference
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    • 2001.10a
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    • pp.14-17
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    • 2001
  • Positional cloning (map-based cloning) of mutations or genetic variations has been served as an invaluable tool to understand in-vivo functions of genes and to identify molecular components underlying phenotypes of interest. Mice homozygous for the cerebellar deficient folia (cdf) mutation are ataxic, with cerebellar hypoplasia and abnormal lobulation of the cerebellum. In the cdf mutant cerebellum approximately 40% of Purkinje cells are ectopically located within the white matter and the inner granule cell layer (IGL). To identify the cdf gene, a high-resolution genetic map for the cdf-gene-encompassing region was constructed using 1997 F2 mice generated from C3H/HeSnJ-cdf/cdf and CAST/Ei intercross. The cdf gene showed complete linkage disequilibrium with three tightly linked markers D6Mit208, D6Mit359, and D6Mit225. A contig using YAC, BAC, and P1 clones was constructed for the cdf critical region to identify the gene. A deletion in the cdf critical region on chromosome 6 that removes approximately 150kb of DNA was identified. A gene associated with this deletion was identified using cDNA selection. cdf mutant mice with the transgenic copy of the identified gene restored the brain abnormalities of the mutant mice. The positional cloning of cdf gene provides a good example showing the identification of a gene could lead to finding a new component of important molecular pathways.

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Cloning and Sequencing of Gene Fragment of Acid Proteinase from Penicillium oxalicum HCLF-34 (Penicillium oxalicum HCLF-34로부터 Acid Proteinase의 부분유전자 Cloning 및 Sequencing)

  • 현성희;천재순;강상순;김진규
    • Korean Journal of Microbiology
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    • v.40 no.1
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    • pp.12-16
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    • 2004
  • Acid proteinase has been discovered in Aspergillus niger (acid protease A) and Cryphonectria parasitica (acid proteinase EapC) and it plays major roles in cheese formation from milk. In this study, a partial gene encoding acid proteinase in Penicillium oxalicum HCLF-34 was cloned by using PCR with degenerate primers corresponding to highly conserved regions of the acid proteinase. The partial acid proteinase gene in P. oxalicum HCLF-34 contains an open reading frame of 438 base pairs and encodes an acid proteinase protein of 146 amino aicds. The predicted amino acid sequences showed 71 % homology with acid protease A and 67% homology with EapC.

Molecular cloning and foreign gene expression of restriction endonuclease fragments of the Hc nuclear polyhedrosis virus DNA (Hc nuclear polyhedrosis virus DNA 제한효소절편의 molecular cloning 과 외래 유전자 발현)

  • Lee, Keun-Kwang
    • Journal of fish pathology
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    • v.8 no.1
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    • pp.31-36
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    • 1995
  • Hc nuclear polyhedrosis virus DNA genome was digested with EcoRI endonuclease, these partial fragments were recombined into the pUC8 plasmid vector and transformed in E. coli JM 83 cell. The genome DNA has 24 EcoRI fragments and 12 fragments of them were subcloned. The four recombinants were named as eNP-O, eNP-Q, eNP-R and eNP-S. The expression of foregin gene of the recombinants was investigated by analysing protein patterns on the SDS-PAGE. The eNP-O, eNP-Q and eNP-R were expressed a different weight of protein as comparision with potypeptide bands of E. coli JM 83 host cell.

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Image Inpainting by Band Matching, Seamless Cloning and Area Sub-Division (밴드 매칭, 경계제거, 영역분할을 이용한 영상 인페인팅)

  • Lee, Su-Bin;Seo, Yong-Duek
    • Journal of Korea Multimedia Society
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    • v.11 no.2
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    • pp.153-162
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    • 2008
  • We propose a novel image inpainting method composed of two parts: band matching and seamless cloning. In band matching, a band enclosing the boundary of a missing region is compared to those from the other parts of the image. The inner area of the minimum difference band is then copied to the missing region. Even though this band matching results in successful inpainting in many practical applications, brightness discontinuity (a seam) may appear between the filled missing region and its neighborhood. We apply seamless cloning to remove such discontinuity between the two regions. However, since this basic method using one patch may not deal with cases where there are abrupt changes of color or brightness along the boundary, we furthermore devise one more step: target sub-division. The target area is subdivided into small sub-areas, and the band matching and seamless cloning are applied to each of them. The multiple results from the sub-division are then ordered according to inpainting quality, which is measured based on the edge map or discontinuity map along the boundary band.

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Cloning and Sequencing of Heterozygous PSS Gene in Pigs (Cloning을 이용한 PSS Hetero 돼지에서의 염기 서열 분석)

  • Yoo J. Y.;Kim G. W.;Lee J. W.;Kim Y. B.;Lee J. Y.;Lee D. H.;Lee H. J.;Yoon J. M.;Park H. Y.
    • Reproductive and Developmental Biology
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    • v.29 no.1
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    • pp.15-18
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    • 2005
  • This study was carried out to analyze the structure of the gene related to porcine stress syndrome (PSS) through cloning from PSS heterozygous pigs and to examine hereditary type associated with PSS. The results obtained from the study were summarized as follows: Amino acid arginine and cysteine exist in dominant gene, N and harmfully recessive gene, n, respectively. It was confirmed that such genes exists as alleles in the sixth chromosome related to PSS. These allelic genes might be inherited according to Mendelian law.

Cloning and Heterologous Expression of Acetyl Xylan Esterase from Aspergillus ficuum

  • Jeong, Hye-Jong;Park, Seung-Mun;Yang, Mun-Sik;Kim, Dae-Hyeok
    • 한국생물공학회:학술대회논문집
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    • 2000.04a
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    • pp.153-156
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    • 2000
  • Xylan, the major hemicellulose component of many plants, occurs naturally in a partially acetylated form and lignin, the most resistant component in plant cell wall degradation, is also attached to ${\beta}-1,4-linked-D-xylose$ backbone through the ester linkage. Esterases are required to release the esterified substituent and acetyl esterases are important in the complete degradation of acetylated polysaccharides, like pectins and xylans. The gene(Axe) encoding acetyl xylan estarase(AXE) was isolated from genomic ${\lambda}$ library from Aspergillus ficuum. Nucleotide sequencing of the Axe gene indicated that the gene was separated with two intervening sequences and the amino acid sequence comparison revealed that it was closely related to that from A. awamori with the 92 % indentity. Heterologous expression of AXE was conducted by using YEp352 and Saccharomyces cerevisae 2805 as a vector and host expression system, respectively. The Axe gene was placed between GAL1 promoter and GAL7 terminator and then this recombinant vector was used to transform S. cerevisiae 2805 strain. Culture filtrate of the transformed yeast was assayed for the presence of AXE activity by spectrophotometry and, comparing with the host strain, four to five times of enzyme activity was detected in culture filtrate of transformed yeast.

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Clonorchis sinensis tropomyosin: Cloning and sequence of partial cDNA amplified by PCR (간흡충 tropomyosin: PCR로 일부분 증폭된 cDNA의 cloning 및 염기서열)

  • 홍성종
    • Parasites, Hosts and Diseases
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    • v.31 no.3
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    • pp.285-292
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    • 1993
  • C. sinensis total RMh was containing large amount of 185 rRNA but little 285 rRNA. The size of the double-stranded cDNA synthesized from poly $(A)^{+}$ mRNA was 0.4-4.2 kb long with tapering unto 9.5 kb. Degenerated oligonucleotides (as 2 sense and 3 antisense Primers) were designed on the conserved regions of the known tropomyosin amino acid sequences. From one out of the PCR amplifications using total CDNA and matrix of primers, a specific gene product, 580 bp in size, was produced. Upon Southern hybridization of the PCR products with Schistosomn mnnsoni tropomyosin (SMTM) CDNA, only one signal appeared at the band of 580 bp product. This 580 bp product was considered to encode C. sinensis tropomyosin (CSTM) and cloned in pGEM-3Zf(-) for DNA sequencing. CSTM cDNA was 575 bp containing one open reading frame of 191 predicted amino acids, which revealed 86.3% homology with SMTM and 51.1% with rrichostronsylur coeubnlormis tropomyosin. CSTM cDNA obtained will serve as a probe in the studies of molecular cloning of CSTM.

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Construction of a Corynebacteriurn glutarnicum-Escherichicr coli Shuttle Vector and Cloning the Homoserine ehydrogenase Gene from C. glutamicum (Corynebacterium glutamicum-Escherichia coli Shuttle Vector 개발과 C.glutamicum 의 Homoserine Dehydrogenase Gene Cloning)

  • 최신건;박종현;신현경
    • Microbiology and Biotechnology Letters
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    • v.19 no.1
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    • pp.31-36
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    • 1991
  • A 7.5 kilobases hybrid plasmid, designated as pCE1301, was constructed by combining Eschurichia cwli plasmid pBELl which carries the kanamycin resistance gene of Tn5 with a cryptic plasmid, pSRl of Corynebacterium glutamicum. pCE1301 was transformed C. glutaicum by PEG-mediated protoplast method and its transformation efficiency was about $3.0\times 10^3$ transformants per $\mu g$ of the hybrid plasmid DNA. The physical map reveals that pCE1301 has single restriction sites for SalI and EcoRl, respectively. 'The kanamycin resistance of pCE1301 was stably maintained in C. glutamicum up to 25 generations and any segregation was not detected. pCI31301 was also introduced into Brevibacterium flavum and E coil, and replicated in those strains. pCE1301 was proved to be useiul in cloning the homoscrine dehydrogenase gene from C. glutamicum.

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A Robust Multiple Set-Top Box Authentication Scheme for IPTV Against Smart Card Cloning Attack (IPTV 환경에서 스마트카드 복제에 강건한 다중 셋톱박스 인증기법)

  • Lim, Ji-Hwan;Oh, Hee-Kuck;Kim, Sang-Jin
    • The KIPS Transactions:PartC
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    • v.17C no.1
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    • pp.37-46
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    • 2010
  • In an IPTV system, the rights of the content and service provider can be protected by using Conditional Access System (CAS) and Digital Right Management (DRM). In case of the CAS, only the authorized user who has structured authentication keys can decrypt the encrypted content. However, since the CAS establishes a secure channel only between content provider and Smart Card (SC), it cannot protect the system against McCormac Hack attack which eavesdrops on unsecure channel between SC and Set-Top Box (STB) and SC cloning attack. In this paper, we propose a robust multi-STB assisted SC / STB authentication protocol which can protect the IPTV system against not only McCormac Hack attack, but also SC cloning attack. The previous works which bind a STB and a SC during the SC registration phase against the SC cloning attack does not support multi-STB environments. The proposed system which dynamically updates the STB information in subscriber management system using the bi-directional channel characteristic of IPTV system can support the user's multi-STB device effectively.