A novel lineage tracing system using dual labeling of fluorescence localization and barcoding

2022 Fiscal Year Final Research Report

• Project/Area Number: 21K19232
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 43:Biology at molecular to cellular levels, and related fields
• Research Institution: Kyushu University
• Principal Investigator: Kawamata Masaki 九州大学, 生体防御医学研究所, 助教 (80602549)
• Project Period (FY): 2021-07-09 – 2023-03-31
• Keywords: Lineage tracing / DNA barcode / CRISPR-Cas9

Outline of Final Research Achievements

Labeling methods for cells have so far only been able to discriminate seveeral colors by fluorescence, and have had the problem of low resolution. Therefore, we constructed an AIMS that can identify the induction of indels by CRISPR-Cas9 for two alleles by fluorescence, and developed a system that can identify 9 patterns by fluorescent localization patterns. In addition, by simultaneously labeling DNA barcodes using indels induced by CRISPR-Cas9, we have developed a system that enables analysis of different clones among the same color. We confirmed that the fluorescent patterns and barcodes are diverse in the cell population, and demonstrated the usefulness of the new dual-labeling lineage tracing system.

Free Research Field

分子生物学、発生学、再生医療

Academic Significance and Societal Importance of the Research Achievements

これまでのlineage tracingシステムは、蛍光のみで見分けられる細胞数が４個程度で解像度が低い解析系となり、実験結果から誤った仮説を提唱する恐れがあった。今回の二重標識技術を利用することで、識別できる細胞数を格段に増やすことができた。解析精度が格段に高まることによって、今まで築くことができなかったような真の生命現象を解き明かすことに貢献できる点で、学術的意義は大きい。