• Title/Summary/Keyword: random sequencing

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Screening Peptides Binding Specifically to Colorectal Cancer Cells from a Phage Random Peptide Library

  • Wang, Jun-Jiang;Liu, Ying;Zheng, Yang;Liao, Kang-Xiong;Lin, Feng;Wu, Cheng-Tang;Cai, Guan-Fu;Yao, Xue-Qing
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.1
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    • pp.377-381
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    • 2012
  • The aim of this study was to screen for polypeptides binding specifically to LoVo human colorectal cancer cells using a phage-displayed peptide library as a targeting vector for colorectal cancer therapy. Human normal colorectal mucous epithelial cells were applied as absorber cells for subtraction biopanning with a c7c phage display peptide library. Positive phage clones were identified by enzyme-linked immunosorbent assay and immunofluorescence detection; amino acid sequences were deduced by DNA sequencing. After 3 rounds of screening, 5 of 20 phage clones screened positive, showing specific binding to LoVo cells and a conserved RPM motif. Specific peptides against colorectal cancer cells could be obtained from a phage display peptide library and may be used as potential vectors for targeting therapy for colorectal cancer.

Analysis and Identification of Expressed Sequence Tags in Hairy Root Induced from Korean Ginseng (Panax ginseng C. A. Meyer)

  • Yang, Deok-Chun;In, Jun-Gyo
    • Korean Journal of Medicinal Crop Science
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    • v.12 no.2
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    • pp.154-162
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    • 2004
  • Hairy roots were induced from Korean ginseng (Panax ginseng C. A. Meyer) root explants and studied for their gene expression. A total of 3,000 ESTs (expressed sequence tags) from ginseng hairy root were determined and about 2,700 ESTs have a length of readable sequence, which result in 1,352 unique ESTs sequences. The 879 ESTs showed significant similarities to known nucleotide or amino acid sequences in other plant species, which were divided into eleven categories depending upon gene function. The remaining 473 sequences showed no significant matches, which are likely to be transcripts or to be matched to other organisms. The results indicated that the analysis of the ginseng hairy root ESTs by partial sequencing of random cDNA clones may be an efficient approach to isolate genes that are functional in ginseng root in a large scale. Our extensive EST analysis of genes expressed in ginseng hairy root not only contributes to the understanding of the dynamics of genome expression patterns in root organ but also adds data to the repertoire of all genomic genes.

Cloning and characterization of Giardia intestinalis cyclophilin

  • Yu, Hak-Sun;Kong, Hyun-Hee;Chung, Dong-Il
    • Parasites, Hosts and Diseases
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    • v.40 no.3
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    • pp.131-138
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    • 2002
  • The cyclophilins (Cyps) are family members of proteins that exhibit peptidylprolyl cis-trans isomerase (PPIase, EC 5.2.1.8) activity and bind the immunosuppressive agent cyclosprin A (CsA) in varying degrees. During the process of random sequencing of a cDNA library made from Giardia intestinalis WB strain, the cyclophilin gene (gicypl) was isolated. An open reading frame of gicyp1 gene was 576 nucleotides, which corresponded to a translation product of 176 amino acids (Gicypl). The identity with other Cyps was about 58-71%. The 13 residues that constituted the CsA binding site of human cyclophilin were also detected in the amino acid sequence of Gicypl, including tryptophan residue essential for the drug binding. The single copy of the gicypl gene was detected in the G. intestinalis chromosome by southern hybridization analysis. Recombinant Gicyp 1 protein clearly accelerated the rate of cis ${\rightarrow}$ trans isomerization of the peptide substrate and the catalysis was completely inhibited by the addition of $0.5{\;}{\mu}M$ CsA.

Arg243, Invariably Critical for the Transcriptional Activation of Yeast Gcn4p

  • Cho, Gyu-Chull;Lee, Jae-Yung;Kim, Joon
    • Journal of Microbiology
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    • v.37 no.3
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    • pp.154-158
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    • 1999
  • The arginine residue at position 243 (Arg 243) of the yeast transcription factor, Gcn4p, is invariably conserved among bZIP transcription factors. Using site-directed oligonucleotide saturation mutagenesis involving two-step polymerase chain reaction (PCR) amplification, random mutations were successfully introduced at the codon of 243 in the basic domain of Gcn4p. This mutant library was transformed ito Gcn4p defective yeast strain and selected for the transcriptionally active colonies. All colonies which were transcriptionally active had arginines in the codon 243. In this study, the strand preference by Taq polymerase during mutagenesis was also tested. Oligonucleotides were specially designed to test whether or not the polymerase was preferred using the strand as a template. A population of randomly mutated products were cloned into an appropriate vector and characterized by DNA sequencing analysis. Saturation mutagenesis which was performed efficiently by this method revealed a strong bias in terms of strand preference of Taq polymerase by an approximate ratio of 3 to 1 in this study.

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DNA 염기 서열의 단편 조립 프로그램 개발

  • Lee, Byung-Uk;Park, Kie-Jung;Park, Wan;Park, Yong-Ha
    • Microbiology and Biotechnology Letters
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    • v.25 no.6
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    • pp.560-565
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    • 1997
  • DNA fragment assembly is a major concem in shot-gun DNA sequencing project. It is to reconstruct a consensus DNA sequence from a collection of random oritented fragments. We developed a computer program that is useful for DNA fragment assembly. Inputs to the program are DNA fragment sequences including IUB-IUPAC bases. The program produces the most probable reconstruction ot the original DNA sequence as a text format or a PostScript format. The program consists of four phases: the first phase quickly eliminates fragment pairs that can not possibly overlap. In the second phase, the quality of overlap between each pair is calculated to a score. In the third phase, overlap pairs are sorted by their scores and consistency of the overlaps is checked. The last phase determines consensus sequences and displays them. The performance of fragment assembly program was tested on a set of DNA fragment sequences which were generated from long DNA sequences of GenBank by a fragmentation program.

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Applications of Transposon-Based Gene Delivery System in Bacteria

  • Choi, Kyoung-Hee;Kim, Kang-Ju
    • Journal of Microbiology and Biotechnology
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    • v.19 no.3
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    • pp.217-228
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    • 2009
  • Mobile genetic segments, or transposons, are also referred to as jumping genes as they can shift from one position in the genome to another, thus inducing a chromosomal mutation. According to the target site-specificity of the transposon during a transposition event, the result is either the insertion of a gene of interest at a specific chromosomal site, or the creation of knockout mutants. The former situation includes the integration of conjugative transposons via site-specific recombination, several transposons preferring a target site of a conserved AT-rich sequence, and Tn7 being site-specifically inserted at attTn7, the downstream of the essential glmS gene. The latter situation is exploited for random mutagenesis in many prokaryotes, including IS (insertion sequence) elements, mariner, Mu, Tn3 derivatives (Tn4430 and Tn917), Tn5, modified Tn7, Tn10, Tn552, and Ty1, enabling a variety of genetic manipulations. Randomly inserted transposons have been previously employed for a variety of applications such as genetic footprinting, gene transcriptional and translational fusion, signature-tagged mutagenesis (STM), DNA or cDNA sequencing, transposon site hybridization (TraSH), and scanning linker mutagenesis (SLM). Therefore, transposon-mediated genetic engineering is a valuable discipline for the study of bacterial physiology and pathogenesis in living hosts.

Molecular Biological Species Identification of Imported Groupers(Epinephelus moara ♀×E. lanceolatus ♂) (수입산 바리과(Family Serranidae) 잡종 어류(Epinephelus moara ♀×E. lanceolatus ♂)의 분자생물학적 판별)

  • Kim, Yong Hwi;Park, Jong Yeon;Kim, Jae Hoon;Bang, In-Chul
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.53 no.4
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    • pp.566-571
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    • 2020
  • To classify a presumed hybrid of imported grouper species acquired from the National Fishery Products Quality Management Service, maternal and paternal lines were identified based on partial sequencing of mitochondrial cytochrome c oxidase 1 (co1) and nuclear recombination activation gene 1 (rag1) genes. The matrilineal species was identified as Epinephleus moara by a partial (760 bp) co1 sequence. Ambiguous sequences with base pairs belonging to E. moara or E. lanceolatus were found in a total of 15 different base pairs in the partial 1,159 bp of the rag1 gene, and the patrilineal species was found to be E. lanceolatus. Therefore, all of the groupers examined in the study were identified to be hybrids of E. moara and E. lanceolatus. In addition, a fast and convenient method using random amplification of polymorphic DNA (RAPD) was established for hybrid discrimination. Hybrids between E. moara ♀ and E. lanceolatus ♂ were identified through specific bands of 387 bp and 433 bp in PRIMER 6.

Molecular Characterization of a Protein Kinase Gene in Chiness Cabbage(Brassica campestrics subsp. napus var. pekinensis)

  • Jeong, Sang-Ho;Ahn, Ji-Hoon;Lee, June-Seung;Lee, Jong-Seob
    • Animal cells and systems
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    • v.1 no.1
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    • pp.135-142
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    • 1997
  • Random sequencing of expressed sequence tags in roots of Chinese cabbage led to isolation of a partial cDNA clone, BR77, which encoded a putative protein kinase. Using the BR77 cDNA as a probe, we isolated a full-length cDNA encoding the Brassica campestris protein kinase 1 (Bcpk1). The Bcpt1 cDNA contained one open reading frame encoding a polypeptide of 439 amino acids. The putative polypeptide consisted of a short N-terminal region and a protein kinase catalytic domain. The catalytic domain of Bcpkl showed a high homology to cAMP- and calcium- phospholipid-dependent subfamilies of serine/threonine protein kineses. Eleven major catalytic domains in protein kineses were well conserved in Bcpk1. However, Bcpk1 contained a unique nonhomologous intervening sequence between subdomains VII and VIII, which was not found in protein kineses of animals and lower eukaryotes. Genomic DNA gel blot analysis showed that Bcpt1 genes might be present as three copies in the Chinese cabbage genome. These imply that Bcpk1 belongs to a plant-specific serine/threonine protein kinase subfamily.

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Development of SCAR Marker for Identification of the Perilla Species

  • Lee, Myoung-Hee;Yang, Ki-Woong;Ha, Tae Joung;Jung, Chan-Sik;Pae, Suk-Bok;Hwang, Chung-Dong;Park, Chang-Hwan;Baek, In-Youl;Kim, Hyeon-Kyeong;Park, Soon-Ki
    • Korean Journal of Breeding Science
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    • v.43 no.4
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    • pp.265-272
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    • 2011
  • This study is to generate SCARs markers for identification of Perilla species. A SCAR is a genomic DNA fragment at a single genetically defined locus that is identified by PCR amplification using a pair of specific oligonucleotide primers. We derived SCARs by sequencing and cloning the both ends of the amplified products of RAPD markers. Sixteen sequence-specific primers were synthesized from eight RAPD markers, which were completely sequenced. We developed the species-specific SCAR markers which could be used successfully in detecting genetic variation in four Perilla species. These markers could be used to verify species-origins of various forms of Perilla germplasms.

A Fragmentation and Search Method of Query Document for Partially Plagiarized Section Detection (부분표절구간 검출을 위한 질의문서의 분할 및 탐색 기법)

  • Ock, Chang-Seok;Seo, Jong-Kyu;Cho, Hwan-Gue
    • Proceedings of the Korea Information Processing Society Conference
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    • 2012.11a
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    • pp.586-589
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    • 2012
  • 표절과 관련된 이슈가 주목받고 있는 상황에서 표절을 검출하는 방법에 대한 연구가 활발히 진행되고 있다. 일반적으로 표절구간 검출을 위해 복잡한 자연어처리와 같은 의미론적 접근방법이 아닌 비교적 단순한 어휘기반의 문자열 처리 방법을 사용한다. 대표적인 방법으로는 지문법 (Fingerprinting)과 서열정렬 (Sequence alignment) 등이 있다. 하지만 이 방법들을 이용하여 대용량 문서에 대한 표절검사를 수행하기에는 시공간적 복잡도의 문제가 발생한다. 본 논문에서는 이러한 단점을 극복하기 위해 NGS (Next Generation Sequencing)에서 사용하는 BWT (Burrows-Wheeler Transform)[1]를 이용한 탐색방법을 응용한다. 또한 부분표절구간을 검출하고 정확도를 향상시키기 위해 질의문서를 분할하여 작은 조각으로 만든 뒤, 조각들에 대한 질의탐색을 수행한다. 본 논문에서는 질의문서를 분할하는 두 가지 방법을 소개한다. 두 가지 방법은 k-mer analysis를 이용한 방법과 random-split analysis를 이용한 방법으로, 각 방법의 장단점을 실험을 통해 분석하고 실제 부분표절구간의 검출 정확도를 측정하였다.