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<p>Cell-tagging strategies with DNA barcodes have enabled the analysis of clone size dynamics and clone-restricted transcriptomic landscapes in heterogeneous populations. However, isolating a target clone that displays a specific phenotype from a complex population remains challenging. Here we present a multi-kingdom genetic barcoding system, CloneSelect, which enables a target cell clone to be triggered to express a reporter gene for isolation through barcode-specific CRISPR base editing. In CloneSelect, cells are first stably tagged with DNA barcodes and propagated so that their subpopulation can be subjected to a given experiment. A clone that shows a phenotype or genotype of interest at a given time can then be isolated from the initial or subsequent cell pools stored during the experiment using CRISPR base editing. CloneSelect is scalable and compatible with single-cell RNA sequencing. We demonstrate the versatility of CloneSelect in human embryonic kidney 293T cells, mouse embryonic stem cells, human pluripotent stem cells, yeast cells and bacterial cells.</p>

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A multi-kingdom genetic barcoding system for precise clone isolation

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年5月21日

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  • Soh Ishiguro,
  • Kana Ishida,
  • Rina C. Sakata,
  • Minori Ichiraku,
  • Ren Takimoto,
  • Rina Yogo,
  • Yusuke Kijima,
  • Hideto Mori,
  • Mamoru Tanaka,
  • Samuel King,
  • Shoko Tarumoto,
  • Taro Tsujimura,
  • Omar Bashth,
  • Nanami Masuyama,
  • Arman Adel,
  • Hiromi Toyoshima,
  • Motoaki Seki,
  • Ju Hee Oh,
  • Anne-Sophie Archambault,
  • Keiji Nishida,
  • Akihiko Kondo,
  • Satoru Kuhara,
  • Hiroyuki Aburatani,
  • Ramon I. Klein Geltink,
  • Takuya Yamamoto,
  • Nika Shakiba,
  • Yasuhiro Takashima,
  • Nozomu Yachie

摘要

Cell-tagging strategies with DNA barcodes have enabled the analysis of clone size dynamics and clone-restricted transcriptomic landscapes in heterogeneous populations. However, isolating a target clone that displays a specific phenotype from a complex population remains challenging. Here we present a multi-kingdom genetic barcoding system, CloneSelect, which enables a target cell clone to be triggered to express a reporter gene for isolation through barcode-specific CRISPR base editing. In CloneSelect, cells are first stably tagged with DNA barcodes and propagated so that their subpopulation can be subjected to a given experiment. A clone that shows a phenotype or genotype of interest at a given time can then be isolated from the initial or subsequent cell pools stored during the experiment using CRISPR base editing. CloneSelect is scalable and compatible with single-cell RNA sequencing. We demonstrate the versatility of CloneSelect in human embryonic kidney 293T cells, mouse embryonic stem cells, human pluripotent stem cells, yeast cells and bacterial cells.