Novel generic technology for genome synthesis derived from Horizontal gene transfer in nature

Research Project

Project/Area Number	18K06196
Research Category	Grant-in-Aid for Scientific Research (C)
Allocation Type	Multi-year Fund
Section	一般
Review Section	Basic Section 43060:System genome science-related
Research Institution	Tokyo Institute of Technology
Principal Investigator	KANEKO Shinya 東京工業大学, 生命理工学院, 助教 (10399694)
Project Period (FY)	2018-04-01 – 2021-03-31
Project Status	Completed (Fiscal Year 2020)
Budget Amount *help	¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000) Fiscal Year 2020: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000) Fiscal Year 2019: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000) Fiscal Year 2018: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Keywords	ゲノム合成 / コンティグDNA / 枯草菌 / 応用微生物 / 形質転換 / 核酸 / ゲノム / バイオテクノロジー / 溶菌 / 長鎖DNA / 培養細胞
Outline of Final Research Achievements	Handling of DNA fragments comprising over several dozens of kb is difficult, because of their fragility when they are in solution. To avoid physical shearing in vitro, assembly of contig DNA fragments have been accomplished within Bacillus subtilis cell using one step protoplast transformation method. Moreover, to avoid physical shearing in the purification step, we demonstrated that large DNA plasmid over 50 kb in size can be introduced into Escherichia coli, or Saccharomyces cerevisiae using the lysate of donor B. subtilis cells. Conjugation protocol has also been established for transfer of large-sized DNA from E. coli to eukaryotic microorganisms (S. cerevisiae). Although transfer to animal culture cell remains, our designed horizontal transfer methods is a versatile procedure for delivery of large-sized DNAs into prokaryotic and eukaryotic microorganisms, which would be a basic platform in the synthetic genome area.
Academic Significance and Societal Importance of the Research Achievements	生物の設計図であるゲノム配列を簡単に解読できる時代に突入した。次の段階としてゲノム（長鎖DNA）を設計・合成し、細胞導入する研究が世界各国で進行している。しかしあまり認識されていないが、DNAは長くなればそれだけ物理的な衝撃に弱く扱いが難しくなる。本研究の成果は簡便に「長鎖DNAを合成する」だけでなく、合成後の「長鎖DNAを如何に目的の宿主に導入するか」を解決する上で重要な基盤技術を提供する。