Development of electrochemical gene detection method using small molecule ligand

2023 Fiscal Year Final Research Report

• Project/Area Number: 19K05534
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 34020:Analytical chemistry-related
• Research Institution: National Defense Medical College
• Principal Investigator: Takei Fumie 防衛医科大学校(医学教育部医学科進学課程及び専門課程、動物実験施設、共同利用研究施設、病院並びに防衛, 進学課程, 准教授 (30252711)
• Project Period (FY): 2019-04-01 – 2024-03-31
• Keywords: DNA / EIS / 電気化学 / 蛍光分子 / RNA

Outline of Final Research Achievements

We carried out experiments to develop a PCR monitoring system using electrochemical impedance spectroscopy (EIS) suitable for PCR with high salt concentration solutions.EIS usually uses [Fe(CN)6]3-/4- as a mediator, but to make this monitoring method even more sensitive A new EIS method using a 2,7-diaminonaphthyridine derivative (DANP), which specifically binds to the cytosine bulge structure of DNA, as a second mediator was investigated to make this monitoring method even more sensitive.
First, the effectiveness of DANP as a mediator was measured. The results showed that the EIS signal was reduced to 40% of that before the addition of DANP, indicating that DANP has sufficient capacity as a second mediator.
Furthermore, DNA with a cytosine bulge structure was detected by the EIS method, and the signal intensity changed significantly with the addition of DNA, suggesting that DNA with a cytosine bulge structure can be adequately detected by this method.

Free Research Field

核酸化学

Academic Significance and Societal Importance of the Research Achievements

電気化学的な遺伝子検出法は、研究室レベルでは様々な方法が発表されているが、その多くはDNAそのもののターゲットDNA認識能、構造変化能を使ったものである。そのため、目的とする遺伝子ごとに検出用DNAプローブの合成・電極上への固定化が必要となり、簡便化されていない。本提案研究ではDNAの特殊構造に特異的に結合する“小分子リガンド”をセカンドメティエータとして使用することで、電極上にプローブ等の固定化をしなくてもDNAの検出ができることを明らかにした。これは今までの方法に比べて、簡単であり今後のDNAの検出技術へと応用でき学術的意義や社会的意義は大きい。