Development of an innovative genome-editing technique using photoactivatible Cas

2022 Fiscal Year Final Research Report

• Project/Area Number: 20K22611
• Research Category: Grant-in-Aid for Research Activity Start-up
• Allocation Type: Multi-year Fund
• Review Section: 0605:Veterinary medical science, animal science, and related fields
• Research Institution: Shiga University of Medical Science
• Principal Investigator: Matsumoto Shoma 滋賀医科大学, 動物生命科学研究センター, 特任助教 (00881517)
• Project Period (FY): 2020-09-11 – 2023-03-31
• Keywords: Genome editing / CRISPR/Cas9 / optogenetics

Outline of Final Research Achievements

Our preliminary experiments revealed that the developmental efficiency of mouse embryos was not suppressed by blue light irradiation. From this result, we have tried to develop an innovative genome-editing technique using photoactivatable Cas9 (paCas9). We succeeded generating piggyBac transposase vector to induce paCas9 into host genome effectively. Although we performed genome editing for mouse embryos using paCas9 mRNA, only wild type genome sequencing was detected. Subsequently, we decided to use paCas9 plasmid to induce target gene knockout for mouse embryos. Genome editing was successfully observed at target loci, however, the efficiencies were quite low. Our results using mouse embryonic stem cells suggested that interior light also induce genome editing by paCas9.

Free Research Field

Developmental engineering

Academic Significance and Societal Importance of the Research Achievements

本研究の成果から、青色光照射条件のさらなる検討や編集効率の改善の必要性が示唆されたが、青色光照射による時空間制御型の遺伝子改変技術が確立することで、ヘテロな細胞集団における特定の遺伝子機能について新たな知見を得ることが可能になる。特に遺伝子改変動物作製においては、相同組換え修復（HDR）活性の高いG2/S期を狙ってゲノム編集を行うことで、目的のゲノム編集を高効率に誘導可能となることが期待される。