Updated on 2024/02/01

写真a

 
Yuki Kasai
 
Organization
Research and Development Initiative Full-time Researcher
Contact information
The inquiry by e-mail is 《here
External link

Degree

  • 博士(理学) ( 神戸大学 )

Education

  • 1992.3
     

    Kobe University   completed

Research History

  • 2012.4 - 2017.3

    Chuo University   Faculty of Science and Engineering

  • 2014.10 - 2015.3

    Ochanomizu University

  • 2013.10 - 2014.3

    Ochanomizu University

  • 2008.4 - 2012.3

    Kitasato University

  • 2010.4 - 2011.3

    Kitasato University   School of Marine Biosciences

  • 2009.6 - 2010.3

    National Institute of Technology and Evaluation

  • 1997.10 - 2008.3

    (株)海洋バイオテクノロジー研究所研究員

  • 1992.4 - 1997.3

    兵庫県立中央農業技術センター生物工学研究所研究員

  • Chuo University   Research Associate

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Professional Memberships

  • 日本生物工学会

  • 日本微生物生態学会

  • 日本ゲノム微生物学会

Research Interests

  • 環境浄化

  • 遺伝子工学

  • 有害物質分解

  • バイオれメディエーション

  • 微細藻類

  • 嫌気ベンゼン分解

Research Areas

  • Life Science / Applied microbiology

  • Life Science / Molecular biology

Papers

  • Precise excision of a selectable marker gene in transgenic Coccomyxa strains by the piggyBac transposase Reviewed

    Yuki Kasai, Kenta Matsuzaki, Fukiko Ikeda, Yuya Yoshimitsu, Shigeaki Harayama

    ALGAL RESEARCH-BIOMASS BIOFUELS AND BIOPRODUCTS   27   152 - 161   2017.11

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:ELSEVIER SCIENCE BV  

    The piggyBac transposon isolated from the cabbage looper moth Trichoplusia ni, is integrated into the host genome, and then excised from it without leaving a footprint. The piggyBac transposon system has been used as a genomic engineering tool in a variety of organisms. In this study, we used two improved versions of the piggyBac transposase (PBase) to create marker-free transgenic strains of the unicellular green alga Coccomyxa sp. strain KJ as follows: Uracil-auxotrophic (Ura(-)) mutants of strain KJ defective in the gene for uridine monophosphate synthase (KJUMPS) were isolated on agar plates containing 5-fluoroorotic acid. Subsequently, cDNA of KJUMPS (cKJUMPS) was cloned between the promoter and terminator of the elongation factor 1 alpha gene to construct a cKJUMPS expression cassette. A DNA fragment carrying the cKJUMPS expression cassette flanked with piggyBac transposon terminal repeats was then constructed (TR_cKJUMPS) and introduced into an Ura-mutant, and Ura(+) transformants were isolated. One of the Ura(+) transformants was named strain TR2-7. Hyperactive PBase (hyPBase) is a mutant PBase with increased excision and integration frequencies. Herein, we synthesized a coding sequence for hyPBase (KJhyPBase), which has optimized codons for expression in strain KJ, and its expression cassette was introduced into strain TR2-7. Fourteen transformants stably carried the KJhyPBase expression cassette, and TR_cKJUMPS was excised from seven of these. We also introduced an expression cassette of KJhyPBase_Ex, which encodes the excision-competent/integration-defective R372A/K375A/D450N mutant of KJhyPBase (KJhyPBase_Ex), into strain TR2-7, and found that the excision frequency of TR_cKJUMPS in KJhyPBase_Ex transformants was significantly higher than that in KJhyPBase transformants. In further experiments, we purified His-tagged KJhyPBase_Ex, and transfected it into strain TR2-7 using electroporation. Under these conditions, TR_cKJUMPS was precisely excised at a frequency of 8.8x10(-8) cell(-1) .The present data extend applications of the present piggyBac transposase-catalyzed excision system in green algae.

    DOI: 10.1016/j.algal.2017.09.007

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  • Stimulation of albino regeneration from rice tissue culture by proline under high osmosis: A possible relationship with an endogenous transposable element os-mudr Reviewed

    S. Yoshida, Y. Kasai, K. Tamaki, K. Watanabe, M. Fujino, C. Nakamura

    Biotechnology and Biotechnological Equipment   12 ( 1 )   3 - 7   1998

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    Proline stimulates proliferation of callus and somaclonal variation, particularly albino regeneration, in rice tissue culture under high osmosis. Albino regenerants from scutellum-derived callus exhibited various deletion patterns in chloroplast DNA, similar to those reported in anther-derived albino regenerants. Through differential display one PCR product was selected which showed enhanced expression by proline in scutellum-derived callus. Northern blot analysis revealed a 2.6 kb major transcript. A deduced amino acid sequence of a partial cDNA clone (976 b) selected from a cDNA library constructed from proline-treated callus showed 36% homology with a maize protein (MURA) encoded by an autonomous regulatory transposable element MuDR. A rice homologue to maize MuDR (designated as Os-MuDR) was shown to be present as a single copy gene in Japanese rice cultivars. Our results suggest a possible relationship between Os-MuDR and the high frequency of somatic mutations including albino regeneration in rice tissue culture. © 1998 Taylor and Francis Group, LLC.

    DOI: 10.1080/13102818.1998.10818956

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Books

  • Microbial Bioremediation of Non-metals: Current Research

    Horizon Scientific Press  2011.6 

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  • Microbial biodegradation: Genomics and Molecular Biology

    Caister Academic Press  2008.1 

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  • 水環境の浄化・改善技術

    シーエムシー出版  2004.11 

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  • Pseudomonas: Volume 3: Biosynthesis of Macromolecules and Molecular Metabolism

    Springer  2004.6 

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MISC

  • Construction of Marker-Free Transgenic Strains of Chlamydomonas reinhardtii Using a Cre/loxP-Mediated Recombinase System

    Yuki Kasai, Shigeaki Harayama

    PLOS ONE   11 ( 8 )   p.e0161733   2016.8

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    Language:English   Publisher:PUBLIC LIBRARY SCIENCE  

    The Escherichia coli bacteriophage P1 encodes a site-specific recombinase called Cre and two 34-bp target sites of Cre recombinase called loxP. The Cre/loxP system has been used to achieve targeted insertion and precise deletion in many animal and plant genomes. The Cre/loxP system has particularly been used for the removal of selectable marker genes to create marker-free transgenic organisms. For the first time, we applied the Cre/loxP-mediated site-specific recombination system to Chlamydomonas reinhardtii to construct markerfree transgenic strains. Specifically, C. reinhardtii strains cc4350 and cc124 carrying an aphVIII expression cassette flanked by two direct repeats of loxP were constructed. Separately, a synthetic Cre recombinase gene (CrCRE), the codons of which were optimized for expression in C. reinhardtii, was synthesized, and a CrCRE expression cassette was introduced into strain cc4350 carrying a single copy of the loxP-flanked aphVIII expression cassette. Among 46 transformants carrying the CrCRE expression cassette stably, the excision of aphVIII by CrCre recombinase was observed only in one transformant. We then constructed an expression cassette of an in-frame fusion of ble to CrCRE via a short linker peptide. The product of ble (Ble) is a bleomycin-binding protein that confers resistance to bleomycin-related antibiotics such as Zeocin and localizes in the nucleus. Therefore, the ble-(linker)-CrCRE fusion protein is expected to localize in the nucleus. When the ble( linker)-CrCRE expression cassette was integrated into the genome of strain cc4350 carrying a single copy of the loxP-flanked aphVIII expression cassette, CrCre recombinase-mediated excision of the aphVIII expression cassette was observed at a frequency higher than that in stable transformants of the CrCRE expression cassette. Similarly, from strain cc124 carrying a single loxP-flanked aphVIII expression cassette, the aphVIII expression cassette was successfully excised after introduction of the ble-(linker)-CrCRE expression cassette. The ble-(linker)-CrCRE expression cassette remained in the genome after excision of the aphVIII expression cassette, and it was subsequently removed by crossing with the wildtype strain. This precise Cre-mediated deletion method applicable to transgenic C. reinhardtii could further increase the potential of this organism for use in basic and applied research.

    DOI: 10.1371/journal.pone.0161733

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  • Construction of a self-cloning system in the unicellular green alga Pseudochoricystis ellipsoidea

    Yuki Kasai, Kohei Oshima, Fukiko Ikeda, Jun Abe, Yuya Yoshimitsu, Shigeaki Harayama

    BIOTECHNOLOGY FOR BIOFUELS   8 ( 94 )   2015.6

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    Language:English   Publisher:BIOMED CENTRAL LTD  

    Background: Microalgae have received considerable interest as a source of biofuel production. The unicellular green alga Pseudochoricystis ellipsoidea (non-validated scientific name) strain Obi appears to be suitable for large-scale cultivation in outdoor open ponds for biodiesel production because it accumulates lipids to more than 30 % of dry cell weight under nitrogen-depleted conditions. It also grows rapidly under acidic conditions at which most protozoan grazers of microalgae may not be tolerant. The lipid productivity of this alga could be improved using genetic engineering techniques; however, genetically modified organisms are the subject of regulation by specific laws. Therefore, the aim of this study was to develop a self-cloning-based positive selection system for the breeding of P. ellipsoidea.
    Results: In this study, uracil auxotrophic mutants were isolated after the mutagenesis of P. ellipsoidea using either ultraviolet light or a transcription activator-like effector nuclease (TALEN) system. The cDNA of the uridine monophosphate synthase gene (PeUMPS) of P. ellipsoidea was cloned downstream of the promoter of either a beta-tubulin gene (PeTUBULIN1) or the gene for the small subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase (PeRBCS) to construct the pUT1 or pUT2 plasmid, respectively. These constructs were introduced into uracil auxotroph strains, and genetically complementary transformants were isolated successfully on minimal agar plates. Use of Noble agar as the solidifying agent was essential to avoid the development of false-positive colonies. It took more than 6 weeks for the formation of colonies of pUT1 transformants, whereas pUT2 transformants formed colonies in 2 weeks. Real-time PCR revealed that there were more PeUMPS transcripts in pUT2 transformants than in pUT1 transformants. Uracil synthesis (Ura(+)) transformants were also obtained using a gene cassette consisting solely of PeUMPS flanked by the PeRBCS promoter and terminator.
    Conclusions: A self-cloning-based positive selection system for the genetic transformation of P. ellipsoidea was developed. Self-cloned P. ellipsoidea strains will require less-stringent containment measures for large-scale outdoor cultivation.

    DOI: 10.1186/s13068-015-0277-0

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  • Tropicibacter naphthalenivorans gen. nov., sp nov., a polycyclic aromatic hydrocarbon-degrading bacterium isolated from Semarang Port in Indonesia

    Theresia Umi Harwati, Yuki Kasai, Yumiko Kodama, Dwi Susilaningsih, Kazuya Watanabe

    INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY   59   392 - 396   2009.2

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    Language:English   Publisher:SOC GENERAL MICROBIOLOGY  

    An aerobic, Gram-negative, motile bacterium, strain C02(T) was isolated from seawater obtained from Semarang Port in Indonesia. Cells of strain C02(T) were peritrichously flagellated and rod-shaped. Strain C02(T) was able to degrade naphthalene, alkylnaphthalenes and phenanthrene. 16S rRNA gene sequence analysis revealed that this strain was affiliated with the family Rhodobacteraceae in the class Alphaproteobacteria and was related most closely to Marinovum algicola FF3(T) (95.7% similarity) and Thalassobius aestuarii JC2049(T) (95.2%). The DNA G + C content of strain C02(T) was 64.6 mol%. The major cellular fatty acids were C(18:1)omega 7c (50.9% of the total), C(16:0) (17.9%), 11 methyl C(18:1)omega 7c (14.7%), C(18:1)omega 9c (2.9%) and C(19:0) cyclo omega 8c (2.4%), and the predominant respiratory lipoquinone was ubiquinone-10. Based on physiological, chemotaxonomic and phylogenetic data, strain C02(T) is suggested to represent a novel species of a new genus, for which the name Tropicibacter naphthalenivorans gen. nov., sp. nov. is proposed. The type strain of Tropicibacter naphthalenivorans is C02(T) (=JCM 14838(T) =DSM 19561(T)).

    DOI: 10.1099/ijs.0.65821-0

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  • Tropicimonas isoalkanivorans gen. nov., sp nov., a branched-alkane-degrading bacterium isolated from Semarang Port in Indonesia

    Theresia Umi Harwati, Yuki Kasai, Yumiko Kodama, Dwi Susilaningsih, Kazuya Watanabe

    INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY   59   388 - 391   2009.2

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    An aerobic, Gram-negative, motile bacterium, strain B51(T), was isolated from seawater obtained from Semarang Port in Indonesia. Cells of strain B51(T) were peritrichously flagellated and rod-shaped. Strain B51(T) was able to degrade alkanes, branched alkanes and alkyl naphthalenes. 16S rRNA gene sequence analysis revealed that strain B51(T) was affiliated with the family Rhodobacteraceae, and was related most closely to Thioclava pacifica TL 2(T) (94.6% similarity). The DNA G + C content of strain B51(T) was 66.5 mol%. The major cellular fatty acids were C(18:1)omega 7c (84.9%), C(18:1)omega 9c (13.8%), C(16:0) (8.7%), C(18:0) (6.4%) and anteiso-C(15:0) (5.8%) and the major quinone was ubiquinone-10. Based on its phenotypic and phylogenetic characteristics, strain B51(T) is considered to represent a novel species of a new genus, for which the name Tropicimonas isoalcanivorans gen. nov., sp. nov. is proposed. The type strain of the type species is B51(T) (=JCM 14837(T) =DSM 19548(T)).

    DOI: 10.1099/ijs.0.65822-0

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  • Characterizing and screening Oil Degrading Microbes for Land and Beach Reclamation in Indonesia

    Dwi Susilaningsih, Theresia Umi Harwati, Yuki Kasai, Kazuya Watanabe, Fumiyoshi Okazaki, Shigeaki Harayama, Yantyati Widyastuti, Bambang Prasetya

    In proceeding of: International Conference on Urbanisation, Land Use, Land Degradation and Environment, At Turkey   2009

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  • Tranquillimonas alkanivorans gen. nov., sp nov., an alkane-degrading bacterium isolated from Semarang Port in Indonesia

    Theresia Umi Harwati, Yuki Kasai, Yumiko Kodama, Dwi Susilaningsih, Kazuya Watanabe

    INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY   58   2118 - 2121   2008.9

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    Strain A34(T), an obligately halophilic, Gram-negative, non-motile, rod-shaped bacterium, was isolated from seawater obtained from Semarang Port in Indonesia. It possesses a pink pigment and degrades short-chain alkanes. It is positive for catalase and oxidase and reduces nitrate to nitrite. Analyses of 16S rRNA gene sequences revealed a clear affiliation of this strain with the family 'Rhodobacteraceae' in the class Alphaproteobacteria, with its closest relatives being Salipiger mucosus A3(T) (94.9% sequence similarity) and Palleronia marisminoris B33(T) (93.4%). The DNA G + C content was 69.1 mol%. The major cellular fatty acids of strain A34(T) were C(18:1)omega 7c (56.2%), C(19:0) cyclo omega 8c (26.0%) and C(16:0) (9.1 %), while the predominant respiratory lipoquinone was ubiquinone-10. Based on the physiological and phylogenetic data, it is proposed that strain A34(T) should be classified in a new genus and species, for which the name Tranquillimonas alkanivorans gen. nov., sp. nov. is proposed. The type strain of Tranquillimonas alkanivorans is strain A34(T) (JCM 14836(T) =DSM 19547(T)).

    DOI: 10.1099/ijs.0.65817-0

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  • Characterization of diverse hydrocarbon-degrading bacteria isolated from Indonesian seawater

    Teresia Umi Harwati, Yuki Kasai, Yumiko Kodama, Dwi Susilaningsih, Kazuya Watanabe

    MICROBES AND ENVIRONMENTS   22 ( 4 )   412 - 415   2007.12

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    Language:English   Publisher:JAPANESE SOC MICROBIAL ECOLOGY, DEPT BIORESOURCE SCIENCE  

    Seawater sampled from the Semarang port in Indonesia was streaked onto inorganic-medium plates coated with weathered crude oil, and 153 strains were isolated. Analyses of 16S rRNA gene sequences identified 67 different phylotypes affiliated with Alphaproteobacteria (111 strains/44 phylotypes), Gammaproteobacteria (8/8) and Actinobacteria (34/15). The organisms represented by 36 phylotypes could transform petroleum components in pure cultures. Many of them were affiliated with genera yet unrelated to hydrocarbon degradation. Strains unaffiliated with known genera were also obtained. Results suggest that many as-yet-unknown hydrocarbon-degrading bacteria are present in tropical marine environments.

    DOI: 10.1264/jsme2.22.412

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  • Degradative capacities and bioaugmentation potential of an anaerobic benzene-degrading bacterium strain DN11

    Yuki Kasai, Yumiko Kodama, Yoh Takahata, Toshihiro Hoaki, Kazuya Watanabe

    ENVIRONMENTAL SCIENCE & TECHNOLOGY   41 ( 17 )   6222 - 6227   2007.9

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    Language:English   Publisher:AMER CHEMICAL SOC  

    Azoarcus sp. strain DN11 is a denitrifying bacterium capable of benzene degradation under anaerobic conditions The present study evaluated strain DN11 for its application to bioaugmentation of benzene-contaminated underground aquifers. Strain DN11 could grow on benzene, toluene, m-xylene, and benzoate as the sole carbon and energy sources under nitrate-reducing conditions, although o- and p-xylenes were transformed in the presence of toluene. Phenol was not utilized under anaerobic conditions. Kinetic analysis of anaerobic benzene degradation estimated its apparent affinity and inhibition constants to be 0.82 and 11 mu M, respectively. Benzene-contaminated groundwater taken from a former coal-distillation plant site was anaerobically incubated in laboratory bottles and supplemented with either inorganic nutrients (nitrogen, phosphorus, and nitrate) alone, or the nutrients plus strain DN11, showing that benzene was significantly degraded only when DN11 was introduced. Denaturing gradient gel electrophoresis of PCR-amplified 16S rRNA gene fragments, and quantitative PCR revealed that DN11 decreased after benzene was degraded. Following the decrease in DN11 16S rRNA gene fragments corresponding to bacteria related to Owenweeksia hongkongensis and Pelotomaculum isophthalicum, appeared as strong bands, suggesting possible metabolic interactions in anaerobic benzene degradation. Results suggest that DN11 is potentially useful for degrading benzene that contaminates underground aquifers at relatively low concentrations.

    DOI: 10.1021/es062842p

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  • Hydroxylations of substituted naphthalenes by Escherichia coli expressing aromatic dihydroxylating dioxygenase genes from polycyclic aromatic hydrocarbon-utilizing marine bacteria

    Kazutoshi Shindo, Ayako Osawa, Yuki Kasai, Nobuko Lba, Ayako Saotome, Norihiko Misawa

    JOURNAL OF MOLECULAR CATALYSIS B-ENZYMATIC   48 ( 3-4 )   77 - 83   2007.9

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    Language:English   Publisher:ELSEVIER SCIENCE BV  

    Bioconversion experiments of various mono- or di-substituted naphthalenes such as dimethylnaphthalenes were carried out using the cells of Escherichia coli that expressed aromatic dihydroxylating dioxygenase genes (phnA1A2A3A4 and phdABCD) from polycyclic aromatic hydrocarbon-utilizing marine bacteria, Nocardioides sp. KP7 and Cycloclasticus sp. A5, respectively. We found that the former dioxygenase PhnA1A2A3A4 had broad substrate preference for these compounds and often was able to hydroxylate their methyl groups. Specifically, 1,4-dimethylnaphthalene was predominantly bioconverted into 1,4-dihydroxymethyl naphthalene. (c) 2007 Elsevier B.V. All rights reserved.

    DOI: 10.1016/j.molcatb.2007.06.007

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  • 嫌気ベンゼン分解菌DN11株を用いる土壌・地下水の浄化技術

    高畑陽, 笠井由紀, 帆秋利洋, 渡辺一哉

    大成建設技術センター報   ( 40 )   43 - 1   2007

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  • Rapid intrinsic biodegradation of benzene, toluene, and xylenes at the boundary of a gasoline-contaminated plume under natural attenuation

    Yoh Takahata, Yuki Kasai, Toshihiro Hoaki, Kazuya Watanabe

    APPLIED MICROBIOLOGY AND BIOTECHNOLOGY   73 ( 3 )   713 - 722   2006.12

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    Language:English   Publisher:SPRINGER  

    A groundwater plume contaminated with gasoline constituents [mainly benzene, toluene, and xylenes (BTX)] had been treated by pumping and aeration for approximately 10 years, and the treatment strategy was recently changed to monitored natural attenuation (MNA). To gain information on the feasibility of using MNA to control the spread of BTX, chemical and microbiological parameters in groundwater samples obtained inside and outside the contaminated plume were measured over the course of 73 weeks. The depletion of electron aceptors (i.e., dissolved oxygen, nitrate, and sulfate) and increase of soluble iron were observed in the contaminated zone. Laboratory incubation tests revealed that groundwater obtained immediately outside the contaminated zone (the boundary zone) exhibited much higher potential for BTX degradation than those in the contaminated zone and in uncontaminated background zones. The boundary zone was a former contaminated area where BTX were no longer detected. Denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction (PCR)-amplified bacterial 16S rRNA gene fragments revealed that DGGE profiles for groundwater samples obtained from the contaminated zone were clustered together and distinct from those from uncontaminated zones. In addition, unique bacterial rRNA types were observed in the boundary zone. These results indicate that the boundary zone in the contaminant plumes served as a natural barrier for preventing the BTX contamination from spreading out.

    DOI: 10.1007/s00253-006-0522-3

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  • RNA-based stable isotope probing and isolation of anaerobic benzene-degrading bacteria from gasoline-contaminated groundwater

    Y Kasai, Y Takahata, M Manefield, K Watanabe

    APPLIED AND ENVIRONMENTAL MICROBIOLOGY   72 ( 5 )   3586 - 3592   2006.5

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    Stable isotope probing (SIP) of benzene-degrading bacteria in gasoline-contaminated groundwater was coupled to denaturing gradient gel electrophoresis (DGGE) of DNA fragments amplified by reverse transcription-PCR from community 16S rRNA molecules. Supplementation of the groundwater with [C-13(6)]benzene together with an electron acceptor (nitrate, sulfate, or oxygen) showed that a phylotype affiliated with the genus Azoarcus specifically appeared in the C-13-RNA fraction only when nitrate was supplemented. This phylotype was also observed as the major band in DGGE analysis of bacteria] 16S rRNA gene fragments amplified by PCR from the gasoline-contaminated groundwater. In order to isolate the Azoarcus strains, the groundwater sample was streaked on agar plates containing nonselective diluted CGY medium, and the DGGE analysis was used to screen colonies formed on the plates. This procedure identified five bacterial isolates (from 60 colonies) that corresponded to the SIP-identified Azoarcus phylotype, among which two strains (designated DN11 and AN9) degraded benzene under denitrifying conditions. Incubation of these strains with [C-14]benzene showed that the labeled carbon was mostly incorporated into (CO2)-C-14 within 14 days. These results indicate that the Azoarcus population was involved in benzene degradation in the gasoline-contaminated groundwater under denitrifying conditions. We suggest that RNA-based SIP identification coupled to phylogenetic screening of nonselective isolates facilitates the isolation of enrichment/isolation-resistant microorganisms with a specific function.

    DOI: 10.1128/AEM.72.5.3586-3592.2006

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  • Development of a PCR method for the detection and quanti cation of benzoyl-CoA reductase genes and its application to monitored natural attenuation

    A Hosoda, Y Kasai, N Hamamura, Y Takahata, K Watanabe

    BIODEGRADATION   16 ( 6 )   591 - 601   2005.12

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    Benzoyl coenzyme A reductase (BCR) catalyzes dearomatization of benzoyl coenzyme A (benzoyl-CoA), which is the central step in the anaerobic degradative pathways for a variety of aromatic compounds. This study developed a PCR method for the detection and quanti cation of BCR genes in bacterial strains and environmental samples. PCR primers were designed by aligning known BCR genes in Thauera, Azoarcus and Rhodopseudomonas species, and their utility was assessed by amplifying BCR fragments from aromatic-hydrocarbon degrading anaerobes and other bacteria. BCR fragments with the expected sizes were obtained from denitrifying and phototrophic aromatics degraders. The positive signals were also obtained from Geobacter metallireducens and xylene-degrading sulfate-reducing bacterium ( strain mXyS1) but not from other aromatics-degrading sulfate-reducing bacteria and aerobic bacteria. When the PCR was used for analyzing a natural attenuation (NA) site, the positive signal was obtained only from gasoline-contaminated groundwater; sequence analysis of these amplicons revealed that most of them exhibited substantial similarities to the known BCRs. Quantitative competitive PCR analysis estimated BCR-gene copies to account for 10-40% of bacterial 16S rRNA gene copies in the contaminated groundwater, indicating that bacteria possessing BCR genes were highly enriched in the contaminated groundwater. In microcosm bioremediation tests using the contaminated groundwater, the copy number of BCR gene was approximately 10-fold increased in the course of aromatics degradation under denitrifying conditions but not under sulfidogenic conditions. These results suggest the utility of the PCR method for assessing the potential of denitrifying bacteria for aromatic-compound degradation in groundwater.

    DOI: 10.1007/s10532-005-0826-5

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  • Physiological and molecular characterization of a microbial community established in unsaturated, petroleum-contaminated soil

    Y Kasai, Y Takahata, T Hoaki, K Watanabe

    ENVIRONMENTAL MICROBIOLOGY   7 ( 6 )   806 - 818   2005.6

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    The microbial communities established in soil samples from an unsaturated, petroleum-contaminated zone and from an adjacent uncontaminated site were characterized by physiological and molecular approaches. Possible electron acceptors such as sulfate and nitrate had been completely depleted in these soil samples. Slurries of these soil samples were incubated in bottles in the presence of hydrocarbon indicators (benzene, toluene, xylene and decane), and the degradation of these compounds was examined. Supplementation with electron acceptors stimulated hydrocarbon degradation, although the stimulatory effect was small in the contaminated soil. The initial degradation rates in the contaminated soil under fermentative/methanogenic conditions were comparable to those under aerobic conditions. The microbial populations in the original soil samples were analysed by cloning and sequencing of polymerase chain reaction (PCR)-amplified bacterial and archaeal 16S rRNA gene fragments, showing that the sequences retrieved from these soils were substantially different. For instance, Epsilonproteobacteria, Gammaproteobacteria, Crenarchaeota and Methanosarcinales could only be detected at significant levels in the contaminated soil. Denaturing gradient gel electrophoresis (DGGE) analyses of 16S rRNA gene fragments amplified by PCR from the incubated soil-slurry samples showed that supplementation of the electron acceptors resulted in a shift in the major populations, while the DGGE profiles after incubating the contaminated soil under the fermentative/methanogenic conditions were not substantially changed. These results suggest that petroleum contamination of the unsaturated zone resulted in the establishment of a fermentative/methanogenic community with substantial hydrocarbon-degrading potential.

    DOI: 10.1111/j.1462-2920.2005.00754.x

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  • Microbial communities in oil-contaminated seawater

    S Harayama, Y Kasai, A Hara

    CURRENT OPINION IN BIOTECHNOLOGY   15 ( 3 )   205 - 214   2004.6

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    Language:English   Publishing type:Book review, literature introduction, etc.   Publisher:CURRENT BIOLOGY LTD  

    Although diverse bacteria capable of degrading petroleum hydrocarbons have been isolated and characterized, the vast majority of hydrocarbon-degrading bacteria, including anaerobes, could remain undiscovered, as a large fraction of bacteria inhabiting marine environments are uncultivable. Using culture-independent rRNA approaches, changes in the structure of microbial communities have been analyzed in marine environments contaminated by a real oil spill and in micro- or mesocosms that mimic such environments. Alcanivorax and Cycloclasticus of the gamma-Proteobacteria were identified as two key organisms with major roles in the degradation of petroleum hydrocarbons. Alcanivorax is responsible for alkane biodegradation, whereas Cycloclasticus degrades various aromatic hydrocarbons. This information will be useful to develop in situ bioremediation strategies for the clean-up of marine oil spills.

    DOI: 10.1016/j.copbio.2004.04.002

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  • ガソリン汚染地下水におけるBTXの自然減衰評価

    高畑陽, 帆秋利洋, 笠井由紀, 渡辺一哉

    大成建設技術センター報   ( 37 )   40 - 1   2004

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  • Molecular characterization and substrate preference of a polycyclic aromatic hydrocarbon dioxygenase from Cycloclasticus sp strain A5

    Y Kasai, K Shindo, S Harayama, N Misawa

    APPLIED AND ENVIRONMENTAL MICROBIOLOGY   69 ( 11 )   6688 - 6697   2003.11

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    Cycloclasticus sp. strain A5 is able to grow with petroleum polycyclic aromatic hydrocarbons (PAHs), including unsubstituted and substituted naphthalenes, dibenzothiophenes, phenanthrenes, and fluorenes. A set of genes responsible for the degradation of petroleum PAHs was isolated by using the ability of the organism to oxidize indole to indigo. This 10.5-kb DNA fragment was sequenced and found to contain 10 open reading frames (ORFs). Seven ORFs showed homology to previously characterized genes for PAH degradation and were designated phn genes, although the sequence and order of these phn genes were significantly different from the sequence and order of the known PAH-degrading genes. The phnA1, phnA2, phnA3, and phnA4 genes, which encode the alpha and beta subunits of an iron-sulfur protein, a ferredoxin, and a ferredoxin reductase, respectively, were identified as the genes coding for PAH dioxygenase. The phnA4A3 gene cluster was located 3.7 kb downstream of the phnA2 gene. PhnA1 and PhnA2 exhibited moderate (less than 62%) sequence identity to the alpha and beta subunits of other aromatic ring-hydroxylating dioxygenases, but motifs such as the Fe(11)binding site and the [2Fe-2S] cluster ligands were conserved. Escherichia coli cells possessing the phnA1A2A3A4 genes were able to convert phenanthrene, naphthalene, and methylnaphthalene in addition to the tricyclic heterocycles dibenzofuran and dibenzothiophene to their hydroxylated forms. Significantly, the E. coli cells also transformed biphenyl and diphenylmethane, which are ordinarily the substrates of biphenyl dioxygenases.

    DOI: 10.1128/AEM.69.11.6688-6697.2003

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  • 27-B-08 ガソリン汚染地下水中のBTX分解菌の解析(バイオレメディエーション,一般講演)

    笠井 由紀, 高畑 陽, 帆秋 利洋, 渡辺 一哉

    日本微生物生態学会講演要旨集   ( 19 )   2003

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  • 総説:石油汚染のバイオレメディエーション

    共著

    土壌環境センター技術ニュース   ( 7 )   73 - 77   2003

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  • Bacteria belonging to the genus Cycloclasticus play a primary role in the degradation of aromatic hydrocarbons released in a marine environment

    Y Kasai, H Kishira, S Harayama

    APPLIED AND ENVIRONMENTAL MICROBIOLOGY   68 ( 11 )   5625 - 5633   2002.11

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    To identify the bacteria that play a major role in the aerobic degradation of petroleum polynuclear aromatic hydrocarbons (PAHs) in a marine environment, bacteria were enriched from seawater by using 2-methylnaphthalene, phenanthrene, or anthracene as a carbon and energy source. We found that members of the genus Cycloclasticus became predominant in the enrichment cultures. The Cycloclasticus strains isolated in this study could grow on crude oil and degraded PAH components of crude oil, including unsubstituted and substituted naphthalenes, dibenzothiophenes, phenanthrenes, and fluorenes. To deduce the role of Cycloclasticus strains in a coastal zone oil spill, propagation of this bacterial group on oil-coated grains of gravel immersed in seawater was investigated in beach-simulating tanks that were I In wide by 1.5 m long by 1 m high. The tanks were two-thirds filled with gravel, and seawater was continuously introduced into the tanks; the water level was varied between 30 cm above and 30 cm below the surface of the gravel layer to simulate a 12-h tidal cycle. The number of Cycloclasticus cells associated with the grains was on the order of 10(3) cells/g of grains before crude oil was added to the tanks and increased to 3 x 10(6) cells/g of grains after crude oil was added. The number increased further after 14 days to 10(8) cells/g of grains when nitrogen and phosphorus fertilizers were added, while the number remained 3 x 10(6) cells/g of grains when no fertilizers were added. PAH degradation proceeded parallel with the growth of Cycloclasticus cells on the surfaces of the oil-polluted grains of gravel. These observations suggest that bacteria belonging to the genus Cycloclasticus play an important role in the degradation of petroleum PAHs in a marine environment.

    DOI: 10.1128/AEM.68.11.5625-5633.2002

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  • A transcriptionally active maize MuDR-like transposable element in rice and its relatives

    N Asakura, C Nakamura, T Ishii, Y Kasai, S Yoshida

    MOLECULAR GENETICS AND GENOMICS   268 ( 3 )   321 - 330   2002.11

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    Two Mu-like transposable elements were cloned from a rice genomic library using a partial cDNA clone that exhibits high homology to the mudrA gene of the maize element MuDR. Database searches led to the identification of six other sequences that carried highly homologous terminal inverted repeats (TIRs). All the rice elements possessed similar to200-by TIRs, and four were flanked by 9-bp target-site duplications (TSDs). The longer of the two cloned elements, OsMu4-2, could potentially encode a protein colinear with a MURA-like transposase, but it had stop codons in the coding region indicating that it is a pseudogene. All the other elements had large internal deletions. Direct dinucleotide repeats were found in two elements at positions flanking the deleted regions, suggesting that the deletions arose via the interrupted-gap-repair mechanism. Sequences related to empty sites of insertion were found in OsMu4-2 and one of the elements identified in the databases. These results provide evidence that the rice OsMu element was active and transposed in the past. Analysis of OsMu4-2 cDNAs revealed two types of transcripts produced by alternative splicing. Genomic Southern analysis suggested that OsMu4-2 was conserved in rice species with the A genome, but a deleted version was unique to japonica subspecies. Some wild rice species harbored paralogous copies of the OsMu element.

    DOI: 10.1007/s00438-002-0737-7

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  • Predominant growth of Alcanivorax strains in oil-contaminated and nutrient-supplemented sea water

    Y Kasai, H Kishira, T Sasaki, K Syutsubo, K Watanabe, S Harayama

    ENVIRONMENTAL MICROBIOLOGY   4 ( 3 )   141 - 147   2002.3

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    We found that bacteria closely related to Alcanivorax became a dominant bacterial population in petroleum-contaminated sea water when nitrogen and phosphorus nutrients were supplied in adequate quantity. The predominance of Alcanivorax bacteria was demonstrated under three experimental conditions: (i) in batch cultures of sea water containing heavy oil; (ii) in columns packed with oil-coated gravel undergoing a continuous sea water flow; and (iii) in a large-scale tidal flux reactor that mimics a beach undergoing tidal cycles with fresh sea water. These results suggest that bacteria related to Alcanivorax are major players in the bioremediation of oil-contaminated marine environments.

    DOI: 10.1046/j.1462-2920.2002.00275.x

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  • The TOL plasmid pWWO xylN gene product from Pseudomonas putida is involved in m-xylene uptake

    Y Kasai, J Inoue, S Harayama

    JOURNAL OF BACTERIOLOGY   183 ( 22 )   6662 - 6666   2001.11

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    The upper operon of the TOL plasmid pWWO of Pseudomonas putida encodes a set of enzymes involved in the conversion of toluene and xylenes to their carboxylic acid derivatives. The last gene of the upper operon, xylN, encodes a 465-amino-acid polypeptide which exhibits significant sequence similarity to FadL, an outer membrane protein involved in fatty acid transport in Escherichia coli. To analyze the role of the xylN gene product, xylN on TOL plasmid pWW0 was disrupted by inserting a kanamycin resistance gene, and the phenotypes of P. putida harboring the wild-type and xylN mutant TOL plasmids were characterized. The growth of P. putida harboring the wild-type TOL plasmid was inhibited by a high concentration of m-xylene, while that of P. putida harboring the xylN mutant TOL plasmid was not. The apparent K, value for the oxidation of m-xylene in intact cells of the xylN mutant was fourfold higher than that of the wild-type strain, although the TOL catabolic enzyme activities in cell extracts from the two strains were almost identical. We therefore presume that the xylN gene product is a porin involved in the transport of m-xylene and its analogues across the outer membrane. Western blot analysis confirmed the localization of xylN in the outer membrane.

    DOI: 10.1128/JB.183.22.6662-6666.2001

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  • 論説:海を守るバクテリア

    生物工学会誌   2001.9

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  • Molecular detection of marine bacterial populations on beaches contaminated by the Nakhodka tanker oil-spill accident

    Y Kasai, H Kishira, K Syutsubo, S Harayama

    ENVIRONMENTAL MICROBIOLOGY   3 ( 4 )   246 - 255   2001.4

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    Language:English   Publisher:BLACKWELL SCIENCE LTD  

    In January 1997, the tanker Nakhodka sank in the Japan Sea, and more than 5000 tons of heavy oil leaked. The released oil contaminated more than 500 km of the coastline, and some still remained even by June 1999, To investigate the long-term influence of the Nakhodka oil spill on marine bacterial populations, sea water and residual oil were sampled from the oil-contaminated zones 10, 18, 22 and 29 months after the accident, and the bacterial populations in these samples were analysed by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments. The dominant DGGE bands were sequenced, and the sequences were compared with those in DNA sequence libraries. Most of the bacteria in the sea water samples were classified as the Cytophaga-Flavobacterium-Bacteroides phylum, a-Proteobacteria or cyanobacteria, The bacteria detected in the oil paste samples were different from those detected in the sea water samples; they were types related to hydrocarbon degraders, exemplified by strains closely related to Sphingomonas subarctica and Alcanivorax borkumensis, The sizes of the major bacterial populations in the oil paste samples ranged from 3.4 x 10(5) to 1.6 x 10(6) bacteria per gram of oil paste, these low numbers explaining the slow rate of natural attenuation.

    DOI: 10.1046/j.1462-2920.2001.00185.x

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  • 論説:流出油のバイオレメディエーション

    笠井 由紀

    ペトロテック   23 ( 5 )   371 - 375   2000.5

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    CiNii Books

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  • Petroleum biodegradation in marine environments

    Shigeaki Harayama, Hideo Kishira, Yuki Kasai, Kazuaki Shutsubo

    Journal of Molecular Microbiology and Biotechnology   1 ( 1 )   63 - 70   1999.8

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    Petroleum-based products are the major source of energy for industry and daily life. Petroleum is also the raw material for many chemical products such as plastics, paints, and cosmetics. The transport of petroleum across the world is frequent, and the amounts of petroleum stocks in developed countries are enormous. Consequently, the potential for oil spills is significant, and research on the fate of petroleum in a marine environment is important to evaluate the environmental threat of oil spills, and to develop biotechnology to cope with them. Crude oil is constituted from thousands of components which are separated into saturates, aromatics, resins and asphaltenes. Upon discharge into the sea, crude oil is subjected to weathering, the process caused by the combined effects of physical, chemical and biological modification. Saturates, especially those of smaller molecular weight, are readily biodegraded in marine environments. Aromatics with one, two or three aromatic rings are also efficiently biodegraded
    however, those with four or more aromatic ring are quite resistant to biodegradation. The asphaltene and resin fractions contain higher molecular weight compounds whose chemical structures have not yet been resolved. The biodegradability of these compounds is not yet known. It is known that the concentrations of available nitrogen and phosphorus in seawater limit the growth and activities of hydrocarbon-degrading microorganisms in a marine environment. In other words, the addition of nitrogen and phosphorus fertilizers to an oil-contaminated marine environment can stimulate the biodegradation of spilled oil. This notion was confirmed in the large-scale operation for bioremediation after the oil spill from the Exxon Valdez in Alaska. Many microorganisms capable of degrading petroleum components have been isolated. However, few of them seem to be important for petroleum biodegradation in natural environments. One group of bacteria belonging to the genus Alcanivorax does become predominant in an oil-contaminated marine environment, especially when nitrogen and phosphorus fertilizers are added to stimulate the growth of endogenous microorganisms.

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  • Proline stimulates albino regeneration from anther- and seed-derived rice callus under high osmosis

    S Yoshida, Y Kasai, K Watanabe, M Fujino

    JOURNAL OF PLANT PHYSIOLOGY   155 ( 1 )   107 - 109   1999.7

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    Language:English   Publisher:GUSTAV FISCHER VERLAG  

    Proline added in subculture medium stimulated albino regeneration from anther- and seed-derived callus in rice (Oryza sativa). This proline effect was synergistic with sorbitol. About one half of albino regenerants from the seed-derived callus showed various patterns of plastid DNA deletions. This suggested that the mechanism of proline-induced albinism in seed callus-derived regenerants is similar to that in anther callus-derived regenerants.

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  • 論説:流出油対策としてのバイオテクノロジー

    高圧ガス   1999.2

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  • Stimulation of albino regeneration from rice tissue culture by proline under high osmosis: A possible relationship with an endogenous transposable element Os-MuDR

    S Yoshida, Y Kasai, K Tamaki, K Watanabe, M Fujino, C Nakamura

    BIOTECHNOLOGY & BIOTECHNOLOGICAL EQUIPMENT   12 ( 1 )   3 - 7   1998

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    Language:English   Publisher:DIAGNOSIS PRESS LTD  

    Proline stimulates proliferation of callus and somaclonal variation, particularly albino regeneration, in rice tissue culture under high osmosis. Albino regenerants from scutellum-derived callus exhibited various deletion patterns in chloroplast DNA, similar to those reported in anther-derived albino regenerants. Through differential display one PCR product was selected which showed enhanced expression by proline in scutellum-derived callus. Northern blot analysis revealed a 2.6 kb major transcript. A deduced amino acid sequence of a partial cDNA clone (976 b) selected from a cDNA library constructed from proline-treated callus showed 36% homology with a maize protein (MURA) encoded by an autonomous regulatory transposable element MuDR. A rice homologue to maize MuDR (designated as Os-MuDR) was shown to be present as a single copy gene in Japanese rice cultivars. Our results suggest a possible relationship between Os-MuDR and the high frequency of somatic mutations including albino regeneration in rice tissue culture.

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  • アグロバクテリウムによるカーネーションの形質転換

    岩井豊通, 笠井由紀

    兵庫県農業技術センター研究報告(農業編)   44 ( 44 )   p.5 - 8   1996

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    Language:Japanese   Publisher:兵庫県立中央農業技術センター  

    CiNii Books

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    Other Link: http://agriknowledge.affrc.go.jp/RN/2010561506

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Presentations

  • Gene expression analysis of aerobic and anaerobic benzene degradative genes of Azoarcus sp. DN11

    日本農芸化学会  2017 

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  • Aerobic and anaerobic benzene degradation pathway of Azoarcus sp. strain DN11

    日本農芸化学会2016年度大会  2016 

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  • Cre/loxP mediated selectable marker gene recycling in green microalgae

    2016 

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  • Azoarcus sp. DN11 株の嫌気的ベンゼン分解機構の解析

    日本農芸化学会  2015 

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  • Chlamydomonas reinhardtiiにおける選択マーカーの繰り返し使用を可能にする遺伝的方法の開発

    第12回クラミドモナス研究会  2015 

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  • 油分生産性の高い微細藻類の創出

    微細藻類バイオマス利用シンポジウム  2015 

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  • Pseudococcomyxa sp. KJ株のセルフクローニングによる油脂高生産株の作成

    微細藻類バイオマス利用シンポジウム  2015 

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  • "Pseudococcomyxa ellipsoidea"における選択マーカーの繰り返し使用を可能にする遺伝的方法の開発

    微細藻類バイオマス利用シンポジウム  2015 

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  • 油脂生産緑藻Pseudococcomyxa sp. KJ株におけるインタクトな細胞へのエレクトロポレーション法の開発

    微細藻類バイオマス利用シンポジウム  2015 

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  • パーティクルガン法を用いた油脂生産緑藻Pseudococcomyxa sp. KJ株の形質転換効率の改良

    微細藻類バイオマス利用シンポジウム  2015 

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  • 油脂生産性微細藻類Pseudococcomyxa ellipsoideaにおけるセルフクローニングシステムの開発

    第7回日本ゲノム微生物学会年会  2013 

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  • ベンゼン分解菌 DN11 株の培養条件によるベンゼン分解能の影響

    土木学会第67回年次学術講演会  2013 

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  • Pseudococcomyxa ellipsoidea obi株のRbcS転写開始点の解析

    微細藻類バイオマス利用国際シンポジウム  2013 

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  • Twards the construction of self-cloning system in Pseudochoricystis ellipsoidea

    微細藻類バイオマス利用国際シンポジウム  2013 

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  • 嫌気ベンゼン分解菌Azoarcus sp. DN11株の特異的検出法

    日本水環境学会講演集/日本水環境学会  2011 

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  • 嫌気ベンゼン分解菌Azoarcus sp. DN11株を用いる石炭ガス製造工場跡地の浄化実証試験

    地下水・土壌汚染とその防止対策に関する研究集会講演集/地下水・土壌汚染とその防止対策に関する研究集会  2011 

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  • 微生物を利用した石油汚染環境浄化

    日本生物工学会 北日本支部シンポジウム  2011 

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  • 嫌気ベンゼン分解菌DN11株を用いたバイオレメディエーションの基礎的検討

    地下水・土壌汚染とその防止対策に関する研究集会講演集/地下水・土壌汚染とその防止対策に関する研究集会  2010 

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  • 嫌気ベンゼン分解菌DN11株を用いたバイオオーグメンテーション技術に対する安全性評価

    日本水環境学会年会講演集/日本水環境学会  2009 

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  • 嫌気ベンゼン分解菌Azoarcus sp. DN11株のゲノム解析

    第3回日本ゲノム微生物学会年会要旨集/日本ゲノム微生物学会  2009 

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  • 嫌気ベンゼン分解菌Azoarcus sp. DN11株の解析

    日本微生物生態学会講演要旨集/日本微生物生態学会  2009 

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  • 嫌気ベンゼン分解菌Azoarcus sp. DN11株のゲノム解析

    第12回微生物アカデミー研究会講演要旨集/北里大学感染制御研究機構  2009 

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  • 嫌気ベンゼン分解菌Azoarcus sp. DN11株を用いた汚染地下水のバイオオーグメンテーション技術の基礎的検討

    日本水環境学会年会講演集/日本水環境学会  2008 

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  • Genome analysis of an anaerobic benzene-degrading bacterium strain DN11

    第23回日本微生物生態学会講演要旨集/日本微生物生態学会  2007 

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  • Isolation and characterization of Azoarcus strains degrading benzene under denitrifying conditions

    11th International Symposium on Microbial Ecology, Book of Abstracts/ISME  2006 

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  • 嫌気BTX分解菌Azoarcus sp. DN11株に関する研究

    第22回日本微生物生態学会講演要旨集/日本微生物生態学会  2006 

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  • ガソリン汚染地下水中の嫌気ベンゼン分解菌のRNA-SIPによる同定と単離

    第21回日本微生物生態学会講演要旨集/日本微生物生態学会  2005 

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  • Assessment of chemical and microbiological signatures during natural attenuation of gasoline-contaminated groundwater

    Proceedings of European symposium on environmental biotechnology/European Symposium on Environmental Biotechnology  2004 

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  • Identification of BTX-degrading populations in gasoline-contaminated gourndwater

    10th International Symposium on Microbial Ecology, Book of abstracts/ISME  2004 

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  • ガソリン汚染地下水中のベンゼン分解菌同定へのRNA-SIPの適用

    第20回日本微生物生態学会講演要旨集/日本微生物生態学会  2004 

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  • 石油汚染土壌の分子生態解析

    日本生物工学会大会講演要旨集/日本生物工学会  2003 

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  • 嫌気芳香族化合物分解系遺伝子の分子モニタリング技術の開発

    第19回日本微生物生態学会大会講演要旨集/日本微生物生態学会  2003 

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  • ガソリン汚染地下水中のBTX分解菌の解析

    第19回日本微生物生態学会大会講演要旨集/日本微生物生態学会  2003 

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  • TGGE法による石油分解微生物群集の解析

    第3回マリンバイオテクノロジー学会大会講演要旨集/マリンバイオテクノロジー学会  1999 

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  • 日本海重油流出事故現場の微生物ポピュレーションの解析(2)

    日本生物工学会大会講演要旨集/日本生物工学会  1999 

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  • 日本海重油流出事故現場の石油分解細菌のポピュレーションの解析

    日本生物工学会東日本支部「生物工学フォーラム」要旨集/日本生物工学会東日本支部  1999 

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  • 日本海重油流出事故現場の微生物ポピュレーションの解析

    日本生物工学会大会講演要旨集/日本生物工学会  1998 

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  • Bioremediationの現状と可能性

    日本海洋学会大会講演要旨集 特殊号:秋季/日本海洋学会  1998 

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  • 石油汚染とバイオレメディエーション

    TECNO OCEAN '98国際シンポジウム論文集/社団法人国際海洋科学技術協会  1998 

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Works

  • バイオマスエネルギー技術研究開発/戦略的次世代バイオマスエネルギー利用技術開発事業(次世代技術開発)/高油脂生産微細藻類の大規模培養と回収および燃料化に関する研究開発

    2013.4 -  

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  • 海洋微生物等を活用した県産素材の高付加価値化

    2011.4 -  

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  • バイオマスエネルギー技術研究開発/戦略的次世代バイオマスエネルギー利用技術事業(次世代技術開発)/油分生産性の優れた微細藻類の育種・改良技術の研究開発

    2011.4 -  

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  • 嫌気ベンゼン分解の分子機構の解明

    2009.4 -  

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  • 生分解・処理メカニズムの解析と制御技術の開発

    2002.4 -  

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Research Projects

  • Creation of lipid-hyper-producing Coccomyxa strains by utilizing transcriptome big data

    Grant number:18K05561  2018.4 - 2022.3

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research  Grant-in-Aid for Scientific Research (C)  Chuo University

    KASAI YUKI

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    Grant amount: \4160000 ( Direct Cost: \3200000 、 Indirect Cost: \960000 )

    Coccomyxa mutants deficient in a DYRK1 accumulate lipids twice as fast as wild type strain. The transcriptome analysis showed that a lot of genes involved in lipid production were induced earlier in the DYRK1 mutants than in wild-type strain. For the breeding of Coccomyxa, DYRK1 gene was disrupted by CRISPR/Cas9-based genome editing method. CNX1G disrupted mutants that is unable to assimilate nitrate as a nitrogen source also created to prevent the leakage to outer environments.

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  • 高油脂生産微細藻類の大規模培養と回収および燃料化に関する研究開発

    2015 -  

    委託研究 

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    Grant type:Competitive

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  • 油分生産性の優れた微細藻類の育種・改良技術の開発

    2012 -  

    委託研究 

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    Grant type:Competitive

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  • Molecular analysis of anaerobic benzene degradation

    Grant number:21580118  2009 - 2011

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research  Grant-in-Aid for Scientific Research (C)  Kitasato University

    KASAI Yuki

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    Grant amount: \4940000 ( Direct Cost: \3800000 、 Indirect Cost: \1140000 )

    Intermediates formed during anaerobic benzene degradation by Azoarcus sp. DN11 was identified by gas chromatography/mass spectrometry. The data support initial methylation of benzene to toluene, followed by transformation to benzoate. Benzene-induced gene expression analysis suggested that there was a new methyltransferase gene near an anaerobic toluene degradation gene cluster. The overexpressed recombinant methyltransferase converted benzene to a metabolite that has the same retention time with toluene.

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  • Biodegradation kinetics of oil in deep sea

    Grant number:19360400  2007 - 2009

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research  Grant-in-Aid for Scientific Research (B)  Chiba Institute of Technology

    SHIBATA Kiyoshi, WATANABE Kazuya, KASAI Yuki, TAKIGUCHI Yasuyuki, WATANABE Kazuya

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    Grant amount: \14690000 ( Direct Cost: \11300000 、 Indirect Cost: \3390000 )

    Experimental study has been conducted to investigate possibility of biodegradation of oil in deep sea. Pseudomonas stutzeri has ability to decompose phenol at sea surface condition, however such decomposition was not observed under 2.0 to 5.0MPa pressure. On the other hand, phenol decomposed faster at high pressure than at atmospheric pressure, when sediment taken from deep sea was mixed. It indicates the possibility of the biodegradation of oil in deep sea.

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  • Biological Decomposition of Oils and Oil Spill Risk from Ship Wreck in Deep Sea

    Grant number:16360444  2004 - 2005

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research  Grant-in-Aid for Scientific Research (B)  National Maritime Research Institute

    SHIBATA Kiyoshi, HARA Shoichi, WATANABE Kazuya, KASAI Yuki

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    Grant amount: \14700000 ( Direct Cost: \14700000 )

    Biodegradation experiments of oil have been conducted at 20MPa and 279 and 298K to simulate ship wrecks in deep sea. About 100m1 of natural sea water, taken at Tokyo bay or Sagami bay, was mixed with 0.4ml of oil for 10 to 75days. Phosphate and Ammonium salts were added to the sea water. In most of the runts either n-decane or n-hexadecane was used as the oil. After a certain time period of the mixing, the degradation rate was determined by comparing the concentration rations of originally added oil to the other n-alkane added as an internal standard substance after the runs. The concentration was determined with GC-MS. The experimental results show fluctuation of the degradation process. In one case, more than 50% of the oil was decomposed in 10 days, but in another case almost no degradation was observed. The change in DNA sequence of the microcosm in the water was analyzed by DGGE-PCR. The band pattern after the degradation experiment was changed from that in the original sea water, and in each experiment the band patterns were different. It has not been successful to identify the microcosm which contributes the degradation of oil in high pressure water.
    A data base for the ship wrecks near Japanese islands has been constructed. The wrecks over 100 tonnage in this about 100 years have been investigated and 1,206 of them are filed in the data base. The data included are name, type, tonnage and cargo of the ships, date, position and cause of the accidents, damage on the ship structure, human damage, depth of the wreck, size, date of built, owner or operator, shipbuilder of the ships and treatment after the accident.

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