Establishment of reverse genetics in medaka

2011 Fiscal Year Final Research Report

• Project/Area Number: 19101002
• Research Category: Grant-in-Aid for Scientific Research (S)
• Allocation Type: Single-year Grants
• Research Field: Risk sciences of radiation/Chemicals
• Research Institution: Osaka University
• Principal Investigator: TODO Takeshi 大阪大学, 大学院・医学系研究科, 教授 (90163948)
• Co-Investigator(Kenkyū-buntansha): MITANI Hiroshi 東京大学, 新領域創成科学研究科, 教授 (70181922) ; YUBA Shunnsuke 独立行政法人産業技術総合研究所, 研究員 (40263248) ; KAWASAKI Takashi 独立行政法人産業技術総合研究所, 研究員 (60356839)
• Project Period (FY): 2007 – 2011
• Keywords: メダカ / 逆遺伝学 / 突然変異 / DNA損傷 / DNA修復

Research Abstract

To develop the system to study gene function at tissue level in medaka, we have established two methods in the present study. 1) Reverse genetics : TILLING(Targeting Induced Local Lesions IN Genome) is a reverse genetic strategy that combines random chemical mutagenesis with high-throughput discovery of induced mutations in the target genes. The method has been applied to a variety of plant and animal species. Screening of induced mutation is the most important step in TILLING. We developed the High Resolution Melting(HRM) assay, evaluated its efficacy for screening ENU-induced mutations in the medaka TILLING library. The HRM assay is fast, cost-effective, and robust in detecting variations in DNA. By HRM assay many nonsense mutations were identified and the phenotypes of these nonsense mutants confirmed their loss-of-function nature. 2) Regulation of gene expression in tissue : Infrared laser. evoked gene operator(IR-LEGO) is a microscope system optimized for heating cells at single cell level. Heat stress induces transcription of genes that are under the control of a heat shock promoter in individual targeted cells. We applied this technique to medaka fish carrying a heat shock promoter driven transgene and have succeeded the control of ectopic gene expression.

Research Products

• [Journal Article] Dmrt1 mutation causes a male-to-female sex reversal after the sex determination by Dmy in the medaka (2011)
  • Author(s): Masuyama H, Yamada M, Kamei Y, Fujiwara-Ishikawa T, Todo T, Nagahama Y, Matsuda M
  • Journal Title: Chromosome Res Volume: 20(1) Pages: 163-76
  • DOI: DOI:10.1007/s10577-011-9264-x
  • Peer Reviewed
• [Journal Article] A rapid and easy method forthe qualitative detection of intracellular deposition of inhaled nanoparticles (2011)
  • Author(s): Nakajima H, Ozaki K, Hongyo T, Narama I, Todo T
  • Journal Title: Nanomedicine Volume: 7 Pages: 881-888
  • DOI: DOI:10.1016/j.nano.2011.02.004
  • Peer Reviewed
• [Journal Article] High-resolution melting curve analysis for rapid detection of mutations in a Medaka TILLING library (2010)
  • Author(s): Ishikawa T, Kamei Y, Otozai S, Kim J, Sato A, Kuwahara Y, Tanaka M, Deguchi T, Inohara H, Tsujimura T, Todo T
  • Journal Title: BMC Mol Biol Volume: 11(1) Pages: 70
  • DOI: DOI:10.1186/1471-2199-11-70
  • Peer Reviewed
• [Journal Article] Identification of a functional medaka heat shock promoter and characterization of its ability to induce exogenous gene expression in medaka in vitro and in vivo (2010)
  • Author(s): Oda S, Mikami S, Urushihara Y, Murata Y, Kamei Y, Deguchi T, Kitano T, Fujimori KE, Yuba S, Todo T, Mitani H
  • Journal Title: Zoolog Sci Volume: 27(5) Pages: 410-5
  • URL: http://dx.doi.org/10.2108/zsj.27.410
  • Peer Reviewed
• [Journal Article] Diatom PtCPF1 is a new cryptochrome/ photolyase family member with DNA repair and transcription regulation activity (2009)
  • Author(s): Coesel S, Mangogna M, Ishikawa T, Heijde M, Rogato A, Finazzi G, Todo T, Bowler C, Falciatore A
  • Journal Title: EMBO Rep Volume: 10(6) Pages: 655-61
  • DOI: DOI:10.1038/embor.2009.59
  • Peer Reviewed
• [Journal Article] Kamei Infrared laser-mediated local gene induction in medaka, zebrafish and Arabidopsis thaliana (2009)
  • Author(s): Deguchi T, Itoh M, Urawa H, Matsumoto T, Nakayama S, Kawasaki T, Kitano T, Oda S, Mitani H, Takahashi T, Todo T, Sato J, Okada K, Hatta K, Yuba S
  • Journal Title: Dev Growth Differ Volume: 51(9) Pages: 769-75
  • DOI: DOI:10.1111/j.1440-169X.2009.01135.x
  • Peer Reviewed
• [Journal Article] Common light signaling pathways controlling DNA repair and circadian clock entrainment in zebrafish (2009)
  • Author(s): Hirayama J, Miyamura N, Uchida Y, Asaoka Y, Honda R, Sawanobori K, Todo T, Yamamoto T, Sassone-Corsi P, Nishina H
  • Journal Title: Cell Cycle Volume: 8(17) Pages: 2794-801
  • URL: http://dx.doi.org/10.4161/cc.8.17.9447
  • Peer Reviewed
• [Journal Article] Iwai S, Panda S, Getzoff ED. Functional motifs in the(6-4) photolyase crystal structure make a comparative framework for DNA repair photolyases and clock cryptochromes (2009)
  • Author(s): Hitomi K, DiTacchio L, Arvai AS, Yamamoto J, Kim ST, Todo T, Tainer JA, Iwai S, Panda S, Getzoff ED
  • Journal Title: Proc Natl Acad Sci USA Volume: 106(17) Pages: 6962-7
  • DOI: DOI:10.1073/pnas.0809180106
  • Peer Reviewed
• [Journal Article] Infrared laser-mediated gene induction in targeted single cells in vivo (2009)
  • Author(s): Kamei Y, Suzuki M, Watanabe K, Fujimori K, Kawasaki T, Deguchi T, Yoneda Y, Todo T, Takagi S, Funatsu T, Yuba S
  • Journal Title: Nature Methods Volume: 6(1) Pages: 79-81
  • DOI: DOI:10.1038/nmeth.1278
  • Peer Reviewed
• [Journal Article] DNA damage-induced ubiquitylation of RFC2 subunit of replication factor C complex (2008)
  • Author(s): Tomida J, Masuda Y, Hiroaki H, Ishikawa T, Song I, Tsurimoto T, Tateishi S, Shiomi T, Kamei Y, Kim J, Kamiya K, Vaziri C, Ohmori H, Todo T
  • Journal Title: J Biol Chem Volume: 283(14) Pages: 9071-9
  • DOI: DOI:10.1074/jbc.M709835200
  • Peer Reviewed
• [Journal Article] Development of a convenient in vitro fertilization method using interspecific hybrids between Oryzias latipes and Oryzias curvinotus (2007)
  • Author(s): Kamei Y, Itou J, Oda S, Masui M, KimJH, Ishikawa T, Yuba S, Kinoshita M, Mitani H, Todo T
  • Journal Title: Dev Growth Differ Volume: 49(9) Pages: 721-30
  • DOI: DOI:0.1111/j.1440-169X.2007.00966.x
  • Peer Reviewed
• [Journal Article] A model for triplet mutation formation based on error-prone translesional DNA synthesis opposite UV photolesions (2007)
  • Author(s): Ikehata H., Ono T., Tanaka K., Todo T
  • Journal Title: DNA Repair Volume: 6(5) Pages: 658-68
  • URL: http://dx.doi.org/10.1016/j.dnarep.2006.12.011
  • Peer Reviewed
• [Presentation] Isolation and character ization of p53, ATM and ATR mutants in medaka fish (2011)
  • Author(s): Tomoko Fujiwara. Ishikawa, Takeshi Todo et al
  • Organizer: 14^<th> International Congress of Radiation Research
  • Place of Presentation: Warszawa, Poland
  • Year and Date: 2011-08-30
• [Presentation] High throughput screening of induced mutations in Medaka TILLING library (2011)
  • Author(s): Tomoko Fujiwara. Ishikawa, Takeshi Todo
  • Organizer: 5^<th> Asia and Oceania Conf erence for Photobiology
  • Place of Presentation: 奈良県奈良市
  • Year and Date: 2011-07-31
• [Presentation] メダカATM, ATR変異体の解析、日本放射線影 (2009)
  • Author(s): 石川智子, 亀井保博、藤堂剛
  • Organizer: 響学会第52回大会
  • Place of Presentation: 広島国際会議場
  • Year and Date: 2009-11-12
• [Presentation] Establishment of Reverse Genetics in Medaka (2008)
  • Author(s): Todo T
  • Organizer: The Fourth Aquatic Animal Models of Human Disease co nference
  • Place of Presentation: North Carolina USA
  • Year and Date: 2008-02-01
• [Presentation] Screening of induced pointmutations in Medaka with TILING (2007)
  • Author(s): Todo T
  • Organizer: 13^<th> International congress of Radiat ion Research
  • Place of Presentation: San Francisco USA
  • Year and Date: 2007-07-10