Generation of synthetic rescue system in murine haploid ES cells.

2014 Fiscal Year Final Research Report

• Project/Area Number: 26640124
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• 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): 2014-04-01 – 2015-03-31
• Keywords: ゲノムワイド関連解析 / ハプロイドES細胞

Outline of Final Research Achievements

We performed experiments to generate a novel synthetic rescue system in murine haploid ES cells. For conditional KO system in haploid ES cells, we first tried to insert tamoxifen- inducible cassette, ERT2-iCre-ERT2 into the Rosa 26 locus. Since homologous targeting in haploid ES cells was very low, we used ZFN for the targeting.
Meanwhile, production of CRISPR-gRNA library has bee reported. The library is able to disrupt both alleles of a gene of interest.
We modified both alleles of Oct4 to floxed ones and we introduced CRISPR-gRNA library to disrupt endogenous genes randomly. After disruption of Oct4 by activation of Cre with tamoxifen. We isolated undifferentiated clones. We are analyzing these clones right now.

Free Research Field

分子生物学、発生工学