Development of gene manipulation for mammalian mitochondrial DNA

2014 Fiscal Year Final Research Report

• Project/Area Number: 25640057
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Laboratory animal science
• Research Institution: Tokyo Metropolitan Institute of Medical Science
• Principal Investigator: YONEKAWA Hiromichi 公益財団法人東京都医学総合研究所, 基盤技術研究センター, 特任研究員 (30142110)
• Co-Investigator(Kenkyū-buntansha): SHITARA Hiroshi 公益財団法人東京都医学総合研究所, 基盤技術研究センター, 主任基盤技術研究職員 (90321885)
• Project Period (FY): 2013-04-01 – 2015-03-31
• Keywords: mtDNA / 遺伝子操作 / 人工ヌクレアーゼ / ミトコンドリア内局在型 / マウス / トランスジェニックマウス / TALEN

Outline of Final Research Achievements

As an extension of genome editing technique, we have developed a technique for direct manipulation of mtDNA in mammalian cells and/or individuals.　We have generated expression vectors containing cDNA encoding TALEN protein with mitochondrial signal sequence in conjunction with HA-tag. By immuno-fluorescent staining, we detected strong signals exclusively restricted to mitochondria, which were stained with mitochondria-specific fluorescent dye, in mammalian cells transformed with the expression vector. These results show that TALEN protein was transported into mitochondria. Then, to investigate whether the mitochondrially-restricted TALEN protein does work directly to mtDNA as we had expected, RNAs for tagged-TALEN were synthesized using these vector as templates, injected into pronucleus of mouse fertilized eggs.
Currently, we are examining the reliability of data so far obtained. The experiment to see if the mitochondrially-restricted TALEN protein did work to mtDNA is in progress.

Free Research Field

哺乳類遺伝学