Development of rapid digital quantitative technology using loop-mediated isothermal amplification

2020 Fiscal Year Final Research Report

• Project/Area Number: 18K05543
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 38050:Food sciences-related
• Research Institution: National Agriculture and Food Research Organization
• Principal Investigator: Takakabatake Reona 国立研究開発法人農業・食品産業技術総合研究機構, 食品研究部門, 上級研究員 (20463466)
• Project Period (FY): 2018-04-01 – 2021-03-31
• Keywords: 定量分析 / LAMP / 簡易迅速

Outline of Final Research Achievements

To develop digital quantitative method using loop-mediated isothermal amplification (LAMP), it is indispensable for a duplex detection system. In current digital PCR technique, TaqMan probes which are labeled with two different fluorescent dyes, has been used to distinct amplification products with different emitted wavelengths. The TaqMan method is inapplicable to LAMP since DNA polymerase with high strand displacement activity has been used in the DNA amplification. Molecular beacons are fluorescent nucleic acid probes with a hairpin structure. We attempted for duplex detection of LAMP amplification products using molecular beacons and succeeded to simultaneously detect two targets consisting of genetically modified (GM) and species-specific endogenous sequences in GM soybean and maize events.

Free Research Field

食品分析

Academic Significance and Societal Importance of the Research Achievements

本研究では、デジタルLAMP定量法の開発に先立ち、異なるLAMP増幅産物の同時検出を試みた。一般的に、LAMP法では、標的ごとに増幅産物を区別することが困難であるが、Molecular Beaconを適用することにより、同時検出が可能であることを示した。本研究により、LAMP法においても多検体の同時検出が可能となり、遺伝子検査の効率化の可能性が広がった。