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BIOLOGICAL AND MOLECULAR ANALYSIS OF INDUCTION MECHANISM FOR SOMACLONAL VARIAION IN TOMATO

Research Project

Project/Area Number 12660050
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field 植物保護
Research InstitutionKINKI UNIVERSITY

Principal Investigator

TOYODA Hideyoshi  KINKI UNIVERSITY, FACULTY OF AGRICULTURE, PROFESSOR, 農学部, 教授 (00150805)

Co-Investigator(Kenkyū-buntansha) NONOMURA Teruo  KINKI UNIVERSITY, FACULTY OF AGRICULTURE, LECTURER, 講師 (30319660)
MATSUDA Yoshinori  KINKI UNIVERSITY, FACULTY OF AGRICULTURE, ASSOCIATE PROFESSOR, 助教授 (30268453)
Project Period (FY) 2000 – 2003
Project Status Completed (Fiscal Year 2003)
Budget Amount *help
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2003: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2002: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 2001: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 2000: ¥1,600,000 (Direct Cost: ¥1,600,000)
KeywordsRetrotransposon / Tomato / Detection of gene expression / Micromanipulation / トリコーム / RT-PCR / 組織培養 / うどんこ病 / 変異 / 抵抗性 / 青枯病
Research Abstract

When leaf segments of a tomato cultivar 'Ponderosa' were inoculated with Agrobacterium rhizogenes MAFF07-20001 carrying the binary vectors pRi and pBI121/sGFP, adventitious roots were developed from calli formed at the edges of the segments. Primordial roots that showed green fluorescence under blue light and elongated vigorously on hormone-free medium without loss of the green fluorescence were obtained. They were easily distinguishable from the non-fluorescing roots on the same segments. Successful integration of the sGFP and rol C genes into the chromosome of tomato roots was confirmed by polymerase chain reaction and Sourhem hybridization. The present method enables us to evaluate the hairy root formation without subculture, isolation and DNA analysis. This method was tested on all of commercial cultivars available in Japan(42 cultivars) and 14 breeding lines of tomato. All but two breeding lines produced the hairy roots. Thus, the present method is useful for hairy root production … More in tomato. RT-PCR was used to detect gene expression in situ in single selected cells from tomato callus aggregates. The cytoplasm from one cell was removed with a micropipette viewed under a light microscope and used directly for RT-PCR, followed by nested PCR. This method of removing cytosolic contents prevented the introduction of genomic DNA into the RT-PCR, and only intron-spliced products were amplified when intron-containing genes were used as PCR targets. In addition, transcription of the intron-free gene was possibly detected by simultaneously tracing the intron-containing and intron-free genes using mixed primers for the targeted genes. The present study indicated that some stimuli-activated genes, such as CHI3 and TLC1-retrotransposon long terminal repeat, were constitutively transcribed in tomato callus cells. A single-cell RT-PCR was conducted to detect gene expression in situ in pinpointed trichome cells of tomato leaves. The cytoplasm was removed with the micropipette using a light microscope and directly used for RT-PCR, followed by nested PCR. Two intron-containing genes, glyceraldehydes 3-phosphate dehydrogenase gene and plasma membrane H^+-ATPas gene were constantly expressed in this tissues and therefore used as the indicator, because of easy detection of shorter-size PCR-products produced by splicing. In addition, the sucking of nucleus-free cellular contents was effective to prevent contamination of genomic DNA led to miss-amplification of corresponding genomic DNA sequences of the intron-less genes in the process of RT-PCR and subsequent nested PCR. Thus, the present technique could be applicable to single trichome cells of tomato leaves for directly detecting their gene expression in response to chemical and physical stimulation. Less

Report

(5 results)
  • 2003 Annual Research Report   Final Research Report Summary
  • 2002 Annual Research Report
  • 2001 Annual Research Report
  • 2000 Annual Research Report
  • Research Products

    (47 results)

All Other

All Publications (47 results)

  • [Publications] Takeshi Sameshima et al.: "Cytological events in tomato leaves inoculated with conidia of Blumeria graminis f sp hordei and Oidium neolycopersici KTP-O1"Journal of General Plant Pathology. 70. 7-10 (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Koichi Kashinoto et al.: "Infectivity of a Japanese isolate of Oidium neolycopersici KTP-01 to a European tomato cultivar resistant to O. neolycopersici"Journal of General Plant Pathology. 69. 406-408 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Wataru Oichi et al.: "Cosecutive monitoring for conidiogenesis by Oidium neolycopersici on tomato leaves with a high-fidelity digital microscope"Journal of General Plant Pathology. (in press). (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Mizuki Wada et al.: "Possible amplification of mature mRNAs in single cells of tomato callus aggregates by direct RT-PCR of cytosolic contents suctioned out with a micropipette"Plant Cell Tissue and Organ Culture. (in press). (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Kazuhisa Fujita et al.: "Powdery mildew pathogens can suppress the chitinase gene expression in duced in detached inner epidermis of barley coleoptile"Plant Cell Reports. (in press). (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Yoshinori Matsuda et al.: "Molecular Discrimination of Different Powdery Mildew Fungi Simultaneously Attacking Plant Leaves by the Phylogenetic Analysis of rDNA Internal Transcribed Spacer Sequences Amplified from Their Individual Propagules by Normal and Nested PCRs"Phytopathology. (in press). (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Yoshinori Matsuda et al.: "Recent Research Developments in Cell Biology"Direct RT-PCR amplification of mature mRNAs in single trichome cells of plant leaves(in press). (2004)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Matsuda Y, Sameshima T, Inoue K, Nonomura T, Kakutani K, Takamatsu S, Toyoda H: "Molecular Discrimination of Different Powdery Mildew Fungi Simultaneously Attacking Plant Leaves by the Phylogenetic Analysis of rDNA Internal Transcribed Spacer Sequences Amplified from Their Individual Propagules by Normal and Nested PCRs"Phytopathology. (in press). (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Fujita K, Matsuda Y, Wada M, Nishimura M, Sameshima T, Hirai Y, Mori K, Nonomura T, Kakutani K, Toyoda H: "Powdery mildew pathogens can suppress the chitinase gene expression induced in detached inner epidermis of barley coleoptile"Plant Cell Re. (in press). (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Wada M, Matsuda Y, Fujita K, Nishimura M, Nonomura T, Kakutani K, Toyoda H: "Possible amplification of mature mRNAs in single cells of tomato callus aggregates by direct RT-PCR of cytosolic contents suctioned out with a micropipette"Plant Cell Tissue Organ Culture. (in press). (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Oichi W, Matsuda Y, Sameshima T, Nonomura T, Kakutani K, Nishimura H, Kusakari S, Toyoda H: "Cosecutive monitoring for conidiogenesis by Oidium neolycopersici on tomato leaves with a high-fidelity digital microscope"Journal of General Plant Pathology. (in press). (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Kashimoto K, Sameshima T, Matsuda Y, Oichi W, Nonomura T, Kakutani K, Nakata K, Kusakari S, Toyoda H: "Infectivity of a Japanese isolate of Oidium neolycopersici KTP-01 to a European tomato cultivar resistant to O.neolycopersici"Journal of General Plant Pathology. 63. 406-408 (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Sameshima T, Kashimoto K, Kida K, Matsuda Y, Nonomura T, Kakutani K, Nakata K, Kusakari S, Toyoda H: "Cytological events in tomato leaves inoculated with conidia of Blumeria graminis f sp hordei and Oidium neolycopersici KTP-01"Journal of General Plant Pathology. 63. 7-10 (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Fukumoto T, Nonomura T, Matsuda Y, Komaki S, Moriura N, Kakutani K, Sawabe A, Toyoda H: "Evaluation for efficient hairy root production assessed by GFP-gene expression in tomato cultivars inoculated with Agrobacterium rhizogenes MAFF07-20001"Plant Biotechnology. 20. 257-261 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Kashimoto K, Matsuda Y, Matsutani K, Sameshima T, Kakutani K, Nonomura T, Okada K, Kusakari S, Nakata K, Takamatsu S, Toyoda H: "Morphological and molecular characterization for a Japanese isolate of tomato powdery mildew Oidium neolycopersici and its host range"Journal of General Plant Pathology. 69. 176-185 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Matsuda Y, Kashimoto K, Takikawa Y, Aikawa R, Nonomura T, Toyoda H: "Occurrence of new powdery mildew on greenhouse tomato cultivars"Journal of General Plant Pathology. 67. 294-298 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Nonomura T, Matsuda Y, Ikegami Y, Morikawa Y, Toyoda H: "Induction of morphologically changed petals from mutagen-treated apical buds of rose and plant regeneration from varied petal-derived calli"Plant Biotechnology. 18. 233-236 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Nonomura T, Matsuda Y, Tsuda M, Uranaka K, Toyoda H: "Susceptibility of commercial tomato cultivars to bacterial wilt in hydroponic system"Journal of General Plant Pathology. 67. 224-227 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Matsuda Y, Nishimura M, Wada M, Nonomura T, Kakutani K, Toyoda H: "Direct RT-PCR amplification of mature mRNAs in single trichome cells of plant leaves"Recent Research Developments in Cell Biology. (in press). (2004)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Matsuda Y, Kashimoto K, Saito R, Iida Y, Takikawa Y, Ikeda S, Shinogi T, Kakutani K, Nonomura T, Toyoda H: "Gene cloning for chitinase and chitosanase and their application in plant protection"Chitin Enzymology. 71-78 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2003 Final Research Report Summary
  • [Publications] Takeshi Sameshima et al.: "Cytological events in tomato leaves inoculated with conidia of Blumeriagraminis f sp hordei and Oidium neolycopersici KTP-01"Journal of General Plant Pathology. 70. 7-10 (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Koichi Kashinoto et al.: "Infectivity of a Japanese isolate of Oidium neolycopersici KTP-01 to a European tomato cultivar resistant to O.neolycopersici"Journal of General Plant Pathology. 69. 406-408 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Wataru Oichi et al.: "Cosecutive monitoring for conidiogenesis by Oidium neolycopersici on tomato leaves with a high-fidelity digital microscope"Journal of General Plant Pathology. (in press). (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Mizuki Wada et al.: "Possible amplification of mature mRNAs in single cells of tomato callus aggregates by direct RT-PCR of cytosolic contents suctioned out with a micropipette"Plant Cell Tissue and Organ Culture. (in press). (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Kazuhisa Fujita et al.: "Powdery mildew pathogens can suppress the chitinase gene expression induced in detached inner epidermis of barley coleoptile"Plant Cell Reports. (in press). (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Yoshinori Matsuda et al.: "Molecular Discrimination of Different Powdery Mildew Fungi Simultaneously Attacking Plant Leaves by the Phylogenetic Analysis of rDNA Internal Transcribed Spacer Sequences Amplified from Their Individual Propagules by Normal and Nested PCRs"Phytopathology. (in press). (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Yoshinori Matsuda et al.: "Direct RT-PCR amplification of mature mRNAs in single trichome cells of plant leaves"Recent Research Developments in Cell Biology. (in press). (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] K.Kashimoto, Y.Matsuda, K.Matsutani, T.Sameshima, K.Kakutani, T.Nonomura, K.Okada, S.Kusakari, K.Nakata, H.Toyoda: "Morphological and molecular characterization for a Japanese isolate of tomato powdery mildew Odium neolycopersici and its host range"Journal of General Plant Pathology. 69・(2). (2003)

    • Related Report
      2002 Annual Research Report
  • [Publications] 角谷晃司, 池田千亜紀, 野々村照雄, 松田克礼, 豊田秀吉: "トマト葉原基への変異誘起剤処理条件ならびに変異葉からの個体再生条件の解析"生物環境調節. 40・(1). 99-105 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Y.Matsuda, K.Kashimoto, Y.Takikawa, R.Aikami, T.Nonomura, H.Toyoda: "Occurrence of new powdery mildew on greenhouse tomato cultivars"Journal of General Plant Pathology. 67・(4). 294-298 (2001)

    • Related Report
      2002 Annual Research Report
  • [Publications] T.Nonomura, Y.Ikegami, Y.Morikawa, Y Matsuda, H.Toyoda: "Induction of morphologically changed petals from mutagen-treated apical buds of rose and plant regeneration from varied petal-derived calli"Plant Biotechnology. 18・(3). 233-236 (2001)

    • Related Report
      2002 Annual Research Report
  • [Publications] T.Nonomura, Y.Matsuda, M.Tsuda, K.Uranaka, H.Toyoda: "Susceptibility of commercial tomato cultivars to bacterial wilt in hydroponic system"Journal of General Plant Pathology. 67・(3). 224-227 (2001)

    • Related Report
      2002 Annual Research Report
  • [Publications] H.Toyoda, Y.matsuda, T.Nonomura, Y.Takikawa, Y.Otsu, H.Mori, K.Kakutani: "Chitin Enzymology"Atec Edizioni. 614 (2001)

    • Related Report
      2002 Annual Research Report
  • [Publications] T.Nonomura, Y.Matsuda, Y.Ikegami, Y.Morikawa, H.Toyoda: "Induction of morphologically changed petals from mutagen-treated apical buds of rose and plant regeneration from varied petal-derived calli"Plant Biotechnology. 18. 233-236 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] T.Nonomura, Y Matsuda, M.Tsuda, K.Uranaka, H.Toyoda: "Susceptibility of commercial tomato cultivars to bacterial wilt in hydroponic system"Journal of General Plant Pathology. 67. 224-227 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] T.Nonomura, Y.Matsuda, M.Takasugi, T.Ohtani, T.Hasegawa, K.Miyajima, T.Hatasa, H.Toyoda: "A monitoring system for green fluorescence protein gene-transformed Fusarium oxysporum in melon seedling"Journal of General Plant Pathology. 67. 273-280 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] T.Nonomura, Y.Matsuda, M.Bingo, M.Onishi, K.Matsuda, S.Harada, H.Toyoda: "Algicidal effect of 3-(3-indolyl) butanoic acid, a control agent of the bacterial wilt pathogen, Ralstonia solanacearum"Crop Protection. 20. 935-939 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Y.Matsuda, K.Kashimoto, R.Aikami, Y.Takikawa, T.Nonomura, H.Toyoda: "Occurence of new powdery mildew on greenhouse tomato cultivars"Journal of General Plant Pathology. 67. 294-298 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] H.Toyoda, Y.Matsuda, T.Nonomura, S.Ouchi: "Plant Disease and Their Control"Chaina Agricultural Scientech Press. 171 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] H.Toyoda, Y.Matsuda, T.Nonomura, K.Kakutani: "Chitin Enzymology"Atec Edizioni. 614 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Y.Matsuda,H.Toyoda,Y.Kato,K.Kakutani,T.Nakanisi,M.Bingo,T.Nonomura,S.Ouchi: "Molecular monitoring of hrpB-disrupted mutant of Ralstonia solanacearum in tomato plants"Jouunal of General Plant Pathology. 66. 59-63 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Y.Matsuda,H.Toyoda,A.Sawabe,K.Maeda,N.Shimizu,N.Fujita,T.Fujita,T.Nonomura,S.Ouchi: "A hairy root culture of melon produces aroma compounds"Journal of Agricultural and Food Chemistry. 48. 1417-1420 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] 野々村照雄,豊田秀吉,松田克礼,石倉孝祐,大内成志: "ホウレンソウ萎ちょう病菌に対するインドール類縁化合物の抑制機構とFusarium oxysporumの分化型間における抗菌活性の差異"関西病虫害研報告. 42. 3-8 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] T.Nonomura,Y.Matsuda,M.Bingo,M.Onishi,K.Matsuda,S.Harada,H.Toyoda: "Algicidal effect of 3-(3-indolyl) butanoic acid as chemical control agent of bacterial wilt pathogen, Ralstonia solanacearum"Crop Protection. (in press). (2001)

    • Related Report
      2000 Annual Research Report
  • [Publications] Y.Matsuda,K.Kashimoto,R.Aikami,Y.Takikawa,T.Nonomura,H.Toyoda: "Evaluation of differential disease severity in commercial cultivars of tomato (I) Different susceptibility of tomato cultivars to powdery mildew"Journal of General Plant Pathology. (in press). (2001)

    • Related Report
      2000 Annual Research Report
  • [Publications] H.Toyoda,Y.Matsuda,Y.Takikawa,H.Mori,Y.Otsu,T.Nonomura: "CHITIN ENZYMOLOGY VOL.3"Atec Edizioni-Grottammare (in press). (2001)

    • Related Report
      2000 Annual Research Report
  • [Publications] H.Toyoda,Y.Matsuda,T.Nonomura,S.Ouchi: "Asian Plant Pathology"China Agricultural Scientech Press (in press). (2001)

    • Related Report
      2000 Annual Research Report

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Published: 2000-04-01   Modified: 2016-04-21  

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