Functional analysis of intergenic regions by contiguous introduction of genomic deletion mutations

• Project/Area Number: 15H04284
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Laboratory animal science
• Research Institution: Osaka University
• Principal Investigator: KOKUBU CHIKARA 大阪大学, 医学系研究科, 准教授 (70379238)
• Research Collaborator: TANAKA Sachiyo 大阪大学, 医学系研究科, 特任研究員
• Project Period (FY): 2015-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥17,030,000 (Direct Cost: ¥13,100,000、Indirect Cost: ¥3,930,000); Fiscal Year 2017: ¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000); Fiscal Year 2016: ¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000); Fiscal Year 2015: ¥7,540,000 (Direct Cost: ¥5,800,000、Indirect Cost: ¥1,740,000)
• Keywords: ゲノム工学 / 疾患モデル / 遺伝学 / ゲノム / ゲノム編集

Outline of Final Research Achievements

In order to enable effective functional analysis of genomic regions ranging from several hundred kilobases to several megabases, this study was focused on development of a new method by combining conventional genome engineering and editing technologies.LoxP elements were transposed in the mouse embryonic stem cell genome, followed by simultaneous induction of site-specific Cre-mediated recombination and CRISPR/Cas-mediated editing.This approach allowed introduction of the intended long-range rearrangements into the genome, but the introduction efficiency has not been significantly improved. The positional combinations of the LoxP sites and CRISPR targets remain to be further optimized.

Research Products

• [Journal Article] Collection of homozygous mutant mouse embryonic stem cells arising from autodiploidization during haploid gene trap mutagenesis. (2018)
  • Author(s): Yamanishi A, Matsuba A, Kondo R, Akamatsu R, Tanaka S, Tokunaga M, Horie K, Kokubu C, Ishida Y, Takeda J.
  • Journal Title: Nucleic Acids Res. Volume: 印刷中 Issue: 10 Pages: e63-e63
  • DOI: 10.1093/nar/gky183
  • Peer Reviewed / Open Access
• [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
• [Presentation] A combination of Bloom gene inactivation and CRISPR/Cas9-mediated DNA cleavage may facilitate construction of a loss-of-heterozygosity library of the mouse genome (2015)
  • Author(s): Kamitani T., Yamanishi A., Yoshimura Y., Kokubu C. and Takeda J.
  • Organizer: Conference on Transposition and Genome Engineering 2015
  • Place of Presentation: 奈良春日野国際フォーラム甍
  • Year and Date: 2015-11-17
  • Int'l Joint Research
• [Presentation] Homozygous mutant mouse embryonic stem cell bank arising from autodiploidization during haploid gene-trap mutagenesis (2015)
  • Author(s): Yamanishi A., Kondo R., Matsuba A., Tokunaga M., Kokubu C., Ishida Y. and Takeda J.
  • Organizer: Conference on Transposition and Genome Engineering 2015
  • Place of Presentation: 奈良春日野国際フォーラム甍
  • Year and Date: 2015-11-17
  • Int'l Joint Research