• Title/Summary/Keyword: Genomic BLAST

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PCR-based identification of Pseudomonas fluorescens in diseased olive flounder, Paralichthys olivaceus, in Jeju Island, South Korea

  • Han, So-Ri;Han, Ho-Seok;Evensen, Oystein;Kim, Sung-Hyun
    • Journal of fish pathology
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    • v.30 no.1
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    • pp.67-70
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    • 2017
  • Pseudomonas is currently causing increasing mortality in farmed olive flounder in Jeju Island. It was previously reported that P. anguilliseptica is the pathogen causing the mortality. It is not known whether other sub-species are involved or not. In this study, P. fluorescens was identified from diseased olive flounder by a PCR-based diagnosis. Based on genomic sequencing and BLAST analysis, 5 out of 6 samples were closer with P. fluorescens than P. anguilliseptica. Our finding suggests that P. fluorescens may be the dominant species causing the disease in farmed olive flounder in Jeju Island, South Korea.

Whole Genome Sequencing and Gene Prediction of Cynodon transvaalensis

  • Sol Ji Lee;Chang soo Kim
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2022.10a
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    • pp.237-237
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    • 2022
  • Cynodon transvaalensis belongs to the warm-season grasses and is one of the economically and ecologically important crops. Cynodon species with high heterozygosity are difficult to assemble, so genome research has not been actively conducted. In this study, hybrid assembly was performed by sequencing with Illumina and PacBio. As a result of the assembly, the number of scaffolds and the length of N50 were 1,392, 928 kb, respectively. The completeness of the assembly was confirmed by BSUCO at 98.3%. In addition, as a result of estimating the size of the assembled genome by K-mer analysis (k=25), it was approximately ~413 Mb. A total of 37,060 cds sequences were annotated in the assembled genome, and their functions were identified through blast. After that, we try to complete the assembled genome into a pseudochromosome-level genome through Hi-C technology. These results will not only help to understand the complex genome composition of african bermudagrass, but also provide a resource for genomic and evolutionary studies of grass and other plant species.

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Molecular cloning of the Arabidopsis gene rat3 that is involving in the Agobacterium-mediated planttransformation (Agrobacterium에 의한 식물형질전환에 관여하는 Arabidopsis RAT3 유전자의 분리와 분석)

  • 남재성;양보경;김도훈;정순재;이영병
    • Journal of Life Science
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    • v.11 no.5
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    • pp.423-431
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    • 2001
  • Genomic and cDNA clones containing the RAT3 gene involving in Agobacterium-mediated plant transformation were identified using plant DNA flanking the righ border of a T-DNA rescued from the rat3 mutant as hy-bridization probe. Two highly homologous cDNA clones were identified; one (RAT3-1) weakly hybridized with the probe whereas another (RAT3-2) strongly hybridized with the probe. Both Rat3-1 and Rat3-2 proteins contain a putative signal peptide for secretion. The deduced molecular weights of encoded proteins are 15 kDa. The results of genomic DNA blot analysis and DNA sequencing indicated that RAT3-1 and RAT3-2 exist as single copy genes and they were arranged side by side with just 600 bp distance between them. RAT3-1 was disrupted by the integration of T-DNA into the 3 untranslated region in rat3 mutant. A BLAST search showed that both RAT3-1 and RAT3-2 proteins have homology with only the C-terminal region of $\beta$-1,3-glucanase homologues from Triticum aestivum and Arabidopsis thaliana. Thses $\beta$-1,3-glucanase homologues contain an unusually long C-terminal region with no sig-nificant homology to other $\beta$-1,3-glucanase.

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The Heavy Metal Tolerant Soil Bacterium Achromobacter sp. AO22 Contains a Unique Copper Homeostasis Locus and Two mer Operons

  • Ng, Shee Ping;Palombo, Enzo A.;Bhave, Mrinal
    • Journal of Microbiology and Biotechnology
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    • v.22 no.6
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    • pp.742-753
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    • 2012
  • Copper-containing compounds are introduced into the environment through agricultural chemicals, mining, and metal industries and cause severe detrimental effects on ecosystems. Certain microorganisms exposed to these stressors exhibit molecular mechanisms to maintain intracellular copper homeostasis and avoid toxicity. We have previously reported that the soil bacterial isolate Achromobacter sp. AO22 is multi-heavy metal tolerant and exhibits a mer operon associated with a Tn21 type transposon. The present study reports that AO22 also hosts a unique cop locus encoding copper homeostasis determinants. The putative cop genes were amplified from the strain AO22 using degenerate primers based on reported cop and pco sequences, and a constructed 10,552 base pair contig (GenBank Accession No. GU929214). BLAST analyses of the sequence revealed a unique cop locus of 10 complete open reading frames, designated copSRABGOFCDK, with unusual separation of copCD from copAB. The promoter areas exhibit two putative cop boxes, and copRS appear to be transcribed divergently from other genes. The putative protein CopA may be a copper oxidase involved in export to the periplasm, CopB is likely extracytoplasmic, CopC may be periplasmic, CopD is cytoplasmic/inner membrane, CopF is a P-type ATPase, and CopG, CopO, and CopK are likely copper chaperones. CopA, B, C, and D exhibit several potential copper ligands and CopS and CopR exhibit features of two-component regulatory systems. Sequences flanking indicate the AO22 cop locus may be present within a genomic island. Achromobacter sp. strain AO22 is thus an ideal candidate for understanding copper homeostasis mechanisms and exploiting them for copper biosensor or biosorption systems.

Comparative Genome Analysis of Rathayibacter tritici NCPPB 1953 with Rathayibacter toxicus Strains Can Facilitate Studies on Mechanisms of Nematode Association and Host Infection

  • Park, Jungwook;Lee, Pyeong An;Lee, Hyun-Hee;Choi, Kihyuck;Lee, Seon-Woo;Seo, Young-Su
    • The Plant Pathology Journal
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    • v.33 no.4
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    • pp.370-381
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    • 2017
  • Rathayibacter tritici, which is a Gram positive, plant pathogenic, non-motile, and rod-shaped bacterium, causes spike blight in wheat and barley. For successful pathogenesis, R. tritici is associated with Anguina tritici, a nematode, which produces seed galls (ear cockles) in certain plant varieties and facilitates spread of infection. Despite significant efforts, little research is available on the mechanism of disease or bacteria-nematode association of this bacterium due to lack of genomic information. Here, we report the first complete genome sequence of R. tritici NCPPB 1953 with diverse features of this strain. The whole genome consists of one circular chromosome of 3,354,681 bp with a GC content of 69.48%. A total of 2,979 genes were predicted, comprising 2,866 protein coding genes and 49 RNA genes. The comparative genomic analyses between R. tritici NCPPB 1953 and R. toxicus strains identified 1,052 specific genes in R. tritici NCPPB 1953. Using the BlastKOALA database, we revealed that the flexible genome of R. tritici NCPPB 1953 is highly enriched in 'Environmental Information Processing' system and metabolic processes for diverse substrates. Furthermore, many specific genes of R. tritici NCPPB 1953 are distributed in substrate-binding proteins for extracellular signals including saccharides, lipids, phosphates, amino acids and metallic cations. These data provides clues on rapid and stable colonization of R. tritici for disease mechanism and nematode association.

Localization of 5,105 Hanwoo (Korean Cattle) BAC Clones on Bovine Chromosomes by the Analysis of BAC End Sequences (BESs) Involving 21,024 Clones

  • Choi, Jae Min;Chae, Sung-Hwa;Kang, Se Won;Choi, Dong-Sik;Lee, Yong Seok;Park, Hong-Seog;Yeo, Jung-Sou;Choi, Inho
    • Asian-Australasian Journal of Animal Sciences
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    • v.20 no.11
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    • pp.1636-1650
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    • 2007
  • As an initial step toward a better understanding of the genome structure of Korean cattle (Hanwoo breed) and initiation of the framework for genomic research in this bovine, the bacterial artificial chromosome (BAC) end sequencing of 21,024 clones was recently completed. Among these clones, BAC End Sequences (BESs) of 20,158 clones with high quality sequences (Phred score ${\geq}20$, average BES equaled 620 bp and totaled 23,585,814 bp), after editing sequencing results by eliminating vector sequences, were used initially to compare sequence homology with the known bovine chromosomal DNA sequence by using BLASTN analysis. Blast analysis of the BESs against the NCBI Genome database for Bos taurus (Build 2.1) indicated that the BESs from 13,201 clones matched bovine contig sequences with significant blast hits (E<$e^{-40}$), including 7,075 single-end hits and 6,126 paired-end hits. Finally, a total of 5,105 clones of the Korean cattle BAC clones with paired-end hits, including 4,053 clones from the primary analysis and 1,052 clones from the secondary analysis, were mapped to the bovine chromosome with very high accuracy.

TP53 Codon 72 Polymorphism and Risk of Acute Leukemia

  • Dunna, Nageswara Rao;Vure, Sugunakar;Sailaja, K.;Surekha, D.;Raghunadharao, D.;Rajappa, Senthil;Vishnupriya, S.
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.1
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    • pp.347-350
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    • 2012
  • TP53 is the mostly commonly mutated gene in many cancers and the P53 tumor suppressor protein is involved in multiple cellular processes, including transcription, DNA repair, genomic stability, senescence, cell cycle control and apoptosis. A common single nucleotide polymorphism located within the proline rich region of TP53 gene at codon 72 in exon 4 encodes either proline or arginine. TP53 Arg 72 is more active than TP53 Pro 72 in inducing apoptosis. The aim of this study was to understand the association of the 72 codon polymorphism with acute leukemia development and prognosis. A total of 288 acute leukemia cases comprising 147 acute lymphocytic leukemia (ALL) and 141 acute myeloid leukemia (AML), as well as 245 controls were recruited for analysis of the TP53 72 polymorphism using PCR-RFLP method. Significant association of homozygous arginine genotype with AML was observed (${\chi}^2$- 133.53; df-2, p < 0.001. When data were analyzed with respect to clinical variables, elevation in mean WBC, blast %, LDH levels and slight reduction in DFS in ALL cases with the arginine genotype was observed. In contrast, AML patients with Pro/Pro had elevated WBC, Blast%, LDH levels with slightly reduced DFS. Our study indicates that Arg/Arg genotype might confer increased risk to development of acute myeloid leukemia.

A Rapid and Simple Method for DNA Preparation of Magnaporthe oryzae from Single Rice Blast Lesions for PCR-Based Molecular Analysis

  • Liying, Dong;Shufang, Liu;Jing, Li;Didier, Tharreau;Pei, Liu;Dayun, Tao;Qinzhong, Yang
    • The Plant Pathology Journal
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    • v.38 no.6
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    • pp.679-684
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    • 2022
  • Rice blast is one of the most destructive diseases of rice worldwide, and the causative agent is the filamentous ascomycete Magnaporthe oryzae. With the successful cloning of more and more avirulence genes from M. oryzae, the direct extraction of M. oryzae genomic DNA from infected rice tissue would be useful alternative for rapid monitoring of changes of avirulence genes without isolation and cultivation of the pathogen. In this study, a fast, low-cost and reliable method for DNA preparation of M. oryzae from a small piece of infected single rice leaf or neck lesion was established. This single step method only required 10 min for DNA preparation and conventional chemical reagents commonly found in the laboratory. The AvrPik and AvrPi9 genes were successfully amplified with the prepared DNA. The expected DNA fragments from 570 bp to 1,139 bp could be amplified even three months after DNA preparation. This method was also suitable for DNA preparation from M. oryzae strains stored on the filter paper. All together these results indicate that the DNA preparation method established in this study is reliable, and could meet the basic needs for polymerase chain reaction-based analysis of M. oryzae.

MOLECULAR CLONING AND SEQUENCE ANALYSIS OF THE GENE FOR THE HEMIN-BINDING PROTEIN FROM Prevotella intermedia (Prevotella intermedia에서의 Hemin 결합 단백질 유전자의 분리 및 염기서열 분석)

  • Kim, Shin;Kim, Sung-Jo
    • Journal of the korean academy of Pediatric Dentistry
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    • v.33 no.2
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    • pp.304-310
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    • 2006
  • Prevotella intermedia is one of the most frequently implicated pathogens in human periodontal disease and has a requirement for hemin for growth. This study has identified a hemin-binding P. intermedia protein by expression of a P. intermedia genomic library in Escherichia coli, a bacterium which does not require or transport exogenous hemin. The genomic library of P. intermedia was constructed into plasmid pUC18, transformed into Escherichia coli strain $DH5{\alpha}$, and screened for recombinant clones using heminbinding activity by plating onto hemin-containing agar. Approximately 5,000 recombinant E. coli colonies were screened onto LB-amp-hemin agar, single clone(pHem1) was exhibited a clearly pigmented phonotype. The 2.5kb insert DNA of pHem1 was determined by restriction enzyme mapping. Southern blot analysis of BamHI, BglII, EcoRI, HindIII and PstI-digested P. intermedia DNA indicated that single copy of the gene was present in the genome. Northern blot analysis revealed that the size of transcript was approximately 1.8 kb. The cloned gene contained a single ORF, consisting of approximately 850-residue amino acids. A BLAST search of the Institute for Genomic Research genes with similar nucleotide sequence revealed no significant similarity It needs further investigation to clarify the mechanisms of heme uptake in P. intermedia.

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Cloning of hadA-like Sigma Factor Gene from Streptomyces coelicolor A3(2) (Streptomyces coelicolor A3(2)에서 hrdA유사 Sigma 인자 유전자의 클로닝)

  • Hahn, Ji-Sook;Cho, Eun-Jung;Roe, Jung-Hye
    • Korean Journal of Microbiology
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    • v.32 no.4
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    • pp.264-270
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    • 1994
  • A gene coding for a novel putative $\sigma$ factor of RNA polymerase has been identified from Streptomyces coelicolor A3(2) using Escherichia coli rpoS gene fragment as a probe. The 486 bp rpoS gene fragment was amplified from E. coli genomic DNA by PCR with two synthetic oligonucleotides, the sequences of which were deduced from the amino acid sequences in the regions 2.3 and 4.2 conserved among various bacterial factors. When E. coli genomic DNA fragments were hybridized with cloned rpoS probe, only one band corresponding to rpoS gene (3.2 kb PvuII fragment or 2.3 kb KpnI fragment) was detected. In S. coelicolor, however, two bands were detected both in PvuII digested DNA and SalI digested DNA. 3.5 kb PvuII fragment which binds the rpoS gene probe was cloned (pMS1) from the sublibrary, and the nucleotide sequences of 1.0 kb BamH'/HincII subclone (pBH2) was partially determined. The nucleotide sequences revealed extensive similarity to other $\sigma$ factor genes of S. coelicolor (hrdA, hrdB, hrdC, hrdD), S. aureofaciens (hrdA, hrdB, hrdC, hrdD), Synechococcus species, Pseudomonas aeruginosa, Stigmatella aurantiaca, and Anabaena species. The nucleotide sequences in regions 1.2 and 4 were compared with the corresponding regions of 5 known ${\sigma}$ factor genes of S. coelicolor by multiple alignment. It turned out that the cloned gene is most closely related to hrdA showing 88% amino acid similarity in region 1.2 and 75% in region 4.

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