A gene expression control system using a genome exchange technology on a mammalian artificial chromosome vector

2017 Fiscal Year Final Research Report

• Project/Area Number: 15K12141
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Life / Health / Medical informatics
• Research Institution: Tottori University
• Principal Investigator: Tetsuya Ohbayashi 鳥取大学, 生命機能研究支援センター, 准教授 (80348804)
• Co-Investigator(Kenkyū-buntansha): 古倉 健嗣 鳥取大学, 医学部, 助教 (30344039) ; 中村 和臣 鳥取大学, 生命機能研究支援センター, プロジェクト研究員 (90598137)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Keywords: 人工染色体ベクター / 遺伝子工学 / 遺伝子改変技術

Outline of Final Research Achievements

Mammalian artificial chromosome (MAC) vectors have several advantages as gene delivery vectors, such as stable and independent maintenance in host cells without integration
Previously, a MAC containing a multi-integrase platform (MI-MAC) was developed to facilitate the transfer of multiple genes into desired cells. To introduce multiple-genes into MI-MAC platform, we developed multiple-gene-loading method by combining multi-integration system-equipped MAC vector and CRISPR-Cas9. Next, we developed multiple expression cassette exchange system via three integrase systems on a MAC vector. We designed three excision cassettes that expressed luciferase, where the luciferase expression could be exchanged to a fluorescent protein by site-specific recombination. The transient expression of an integrase caused the targeted luciferase activity to be lost and fluorescence was activated. Furthermore, we developed in vitro cisplatin nephrotoxicity test using these systems.

Free Research Field

実験動物学