Screening drug resistance suppression targets through comprehensive construction of E. coli double knockout strains

2023 Fiscal Year Final Research Report

• Project/Area Number: 20K15779
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 43060:System genome science-related
• Research Institution: Institute of Physical and Chemical Research (2021-2023); Nara Institute of Science and Technology (2020)
• Principal Investigator: Muto Ai 国立研究開発法人理化学研究所, 生命機能科学研究センター, 学振特別研究員RPD (80730506)
• Project Period (FY): 2020-04-01 – 2024-03-31
• Keywords: 薬剤耐性 / 大腸菌 / 網羅的遺伝子欠失株ライブラリ

Outline of Final Research Achievements

In this study, we aimed to identify target genes that suppress bacterial antimicrobial resistance through a comprehensive screening using an E. coli knockout library. We obtained time-series dynamics data by detecting changes in the frequency of each deletion strain, using identifier sequences each strain carries, when the mixed culture of the E. coli gene deletion library was exposed to antibiotics. From this data, we identified candidate drug-resistant strains that showed growth while most of the population died. We then constructed conjugation-transfer strains to comprehensively introduce the gene deletion regions of these candidate strains into the entire gene deletion library. Although we conducted double deletion strain construction experiments on agar plates, the conjugation efficiency of the candidate strains was very low, and we were unable to identify targets for drug resistance suppression.

Free Research Field

システム生物学

Academic Significance and Societal Importance of the Research Achievements

本研究で得られた薬剤下時系列ダイナミクスデータは、薬剤効果の持続性に遺伝子欠失が与える影響を網羅的に検出したものであり、薬剤耐性だけでなく、薬剤への感受性や薬効の継続メカニズムをターゲットとしたスクリーニングにも活用可能である。また、本研究の副産物として得られた自動分注ワークステーションを用いた寒天培地上でのコロニー整列の系は、従来は液体培地に特化していた実験の寒天培地での実施を同一の機器で可能とするものであり、実験自動化の系に組み込むことで相乗効果が期待される。