Identification of genes responsible for disease in human iPS cells

• Project/Area Number: 24241064
• Research Category: Grant-in-Aid for Scientific Research (A)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Medical genome science
• Research Institution: Osaka University
• Principal Investigator: Takeda Junji 大阪大学, 医学(系)研究科(研究院), 教授 (50163407)
• Project Period (FY): 2012-04-01 – 2016-03-31
• Project Status: Completed (Fiscal Year 2015)
• Budget Amount: ¥46,930,000 (Direct Cost: ¥36,100,000、Indirect Cost: ¥10,830,000); Fiscal Year 2014: ¥15,860,000 (Direct Cost: ¥12,200,000、Indirect Cost: ¥3,660,000); Fiscal Year 2013: ¥16,120,000 (Direct Cost: ¥12,400,000、Indirect Cost: ¥3,720,000); Fiscal Year 2012: ¥14,950,000 (Direct Cost: ¥11,500,000、Indirect Cost: ¥3,450,000)
• Keywords: 疾患関連遺伝子 / ヒトiPS細胞

Outline of Final Research Achievements

The purpose of this project is establishment of efficient system for conversion of mono-allelic mutation to bi-allelic mutations in human iPS cell lines. The system is able to identify hidden mutations in human genome without mating.
At first, we made a human iPS cell line in which expression of Bloom syndrome gene (BLM) is suppressed with an addition of doxycycline. Deficiency of BLM results in enhancement of chromosomal crossing over. Combination of BLM deficiency and CRISPR/Cas9-mediated DNA double strand break facilitate site-specific chromosomal crossing over. These results indicate that heterozygous hidden mutations in human genome can be converted to homozygous mutations and identified by mapping of homozygousity.

Research Products

• [Journal Article] Report on the Conference on Transposition and Genome Engineering 2015 (TGE 2015): advancing cutting-edge genomics technology in the ancient city of Nara. (2016)
  • Author(s): Woltjen K, Yamamoto T, Kokubu C, Takeda J.
  • Journal Title: Genes Cells. Volume: - Issue: 5 Pages: 392-395
  • DOI: 10.1111/gtc.12367
  • Peer Reviewed / Open Access
• [Journal Article] Removal of reprogramming transgenes improves the tissue reconstitution potential of keratinocytes generated from human induced pluripotent stem cells (2014)
  • Author(s): Ken Igawaa, Chikara Kokubu, Kosuke Yusa, Kyoji Horie, Yasuhide Yoshimura, Kaori Yamauchi, Hirofumi Suemori, Hiroo Yokozeki, Masashi Toyoda, Nobutaka Kiyokawa, Hajime Okita, Yoshitaka Miyagawa, Hidenori Akutsu, Akihiro Umezawa, Ichiro Katayama, Junji Takeda.
  • Journal Title: Stem Cells Transl Med Volume: 3 Issue: 9 Pages: 992-1001
  • DOI: 10.5966/sctm.2013-0179
  • Peer Reviewed / Open Access
• [Journal Article] Enhancement of microhomology-mediated genomic rearrangements by transient loss of mouse Bloom syndrome helicase. (2013)
  • Author(s): Yamanishi A., Yusa K., Horie K., Tokunaga M., Kusano K., Kokubu C. and Takeda J
  • Journal Title: Genome Research Volume: 23 Issue: 9 Pages: 1462-1473
  • DOI: 10.1101/gr.152744.112
  • Peer Reviewed / Open Access
• [Presentation] A combination of Bloom gene inactivation and CRISPR/Cas9-mediated DNA cleavage facilitates construction of a loss-of-heterozygosity library of the mammalian genome (2015)
  • Author(s): Tomo Kamitani, Ayako Yamanishi, Yasuhide Yoshimura, Chikara Kokubu, Junji Takeda
  • Organizer: Conference on Transpositon and Genome Engineering 2015
  • Place of Presentation: 奈良春日野国際フォーラム 甍～I・RA・KA～（奈良県奈良市）
  • Year and Date: 2015-11-17
  • Int'l Joint Research
• [Presentation] Establishiment of a forward genetic system with chromosomal crossing over in human iPS cells (2015)
  • Author(s): Yasuhide Yoshimura, Tomo Kamitani, Junji Takeda
  • Organizer: Conference on Transpositon and Genome Engineering 2015
  • Place of Presentation: 奈良春日野国際フォーラム 甍～I・RA・KA～（奈良県奈良市）
  • Year and Date: 2015-11-17
  • Int'l Joint Research
• [Presentation] A combination of Bloom gene inactivation and CRISPR/Cas9-mediated DNA cleavage facilitates construction of a loss-of-heterozygosity library of the mammalian genome (2015)
  • Author(s): Junji Takeda, Tomo Kamitani, Ayako Yamanishi, Yasuhide Yoshimura, Chikara Kokubu
  • Organizer: Cold Spring Harbor Laboratory Meeting: Genome Engineering: The Crispr/Cas Revolution
  • Place of Presentation: Cold Spring Harbor Laboratory, (Cold Spring Harbor, USA)
  • Year and Date: 2015-09-24
  • Int'l Joint Research
• [Presentation] ヒトiPS細胞での相同組換え誘発による新規ゲノム解析方法の確立 (2014)
  • Author(s): 吉村 康秀、神谷 智、竹田 潤二
  • Organizer: 第37回日本分子生物学会年会
  • Place of Presentation: パシフィコ横浜（神奈川県横浜市）
  • Year and Date: 2014-11-25
• [Presentation] ES細胞は、遺伝子改変マウス作製に必要不可欠の存在か？ (2013)
  • Author(s): 竹田 潤二
  • Organizer: 第32回日本認知症学会学術集会
  • Place of Presentation: 松本市、日本
  • Year and Date: 2013-11-09
  • Invited
• [Presentation] Gene discovery by transposons in diploid and haploid ES cells (2013)
  • Author(s): 竹田 潤二
  • Organizer: Conference on Transpositoin and Genome Engineering 2013
  • Place of Presentation: Budapest, Hungary
  • Year and Date: 2013-09-19
  • Invited
• [Presentation] マウス一倍体ES細胞を用いたGPIアンカー生合成にかかわる遺伝子の網羅的検索
  • Author(s): 徳永 正浩
  • Organizer: 第35回日本分生生物学会年会
  • Place of Presentation: 福岡国際会議場（福岡県）
• [Remarks] 所属研究室HP
  • URL: http://www.med.osaka-u.ac.jp/pub/mr-envi/www/index.html
• [Remarks] 大阪大学 医学系研究科 環境・生体機能学
  • URL: http://www.med.osaka-u.ac.jp/pub/mr-envi/www/index.html