Letter | Published:

Simultaneous lineage tracing and cell-type identification using CRISPR–Cas9-induced genetic scars

Nature Biotechnology volume 36, pages 469473 (2018) | Download Citation

Abstract

A key goal of developmental biology is to understand how a single cell is transformed into a full-grown organism comprising many different cell types. Single-cell RNA-sequencing (scRNA-seq) is commonly used to identify cell types in a tissue or organ1. However, organizing the resulting taxonomy of cell types into lineage trees to understand the developmental origin of cells remains challenging. Here we present LINNAEUS (lineage tracing by nuclease-activated editing of ubiquitous sequences)—a strategy for simultaneous lineage tracing and transcriptome profiling in thousands of single cells. By combining scRNA-seq with computational analysis of lineage barcodes, generated by genome editing of transgenic reporter genes, we reconstruct developmental lineage trees in zebrafish larvae, and in heart, liver, pancreas, and telencephalon of adult fish. LINNAEUS provides a systematic approach for tracing the origin of novel cell types, or known cell types under different conditions.

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Acknowledgements

We thank R. Opitz, M. Guedes Simoes, D. Panakova, T. Durovic, and J. Ninkovic for help with cell-type identification. We also acknowledge support by MDC/BIMSB core facilities (zebrafish, genomics, bioinformatics), and we thank J. Richter for help with zebrafish experiments. Work in J.P.J.'s laboratory was funded by a European Research Council Starting Grant (ERC-StG 715361 SPACEVAR), a Fondation Leducq Transatlantic Networks Grant (16CVD03), and a Helmholtz Incubator grant (Sparse2Big ZT-I-0007). B.H. was supported by a PhD fellowship from Studienstiftung des deutschen Volkes.

Author information

Author notes

    • Bastiaan Spanjaard
    •  & Bo Hu

    These authors contributed equally to this work.

Affiliations

  1. Berlin Institute for Medical Systems Biology, Max Delbrück Center for Molecular Medicine, Berlin, Germany.

    • Bastiaan Spanjaard
    • , Bo Hu
    • , Nina Mitic
    • , Pedro Olivares-Chauvet
    •  & Jan Philipp Junker
  2. DFG-Center for Regenerative Therapies Dresden, Technische Universität Dresden, Dresden, Germany.

    • Sharan Janjuha
    •  & Nikolay Ninov

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Contributions

J.P.J., B.S. and B.H. conceived and designed the project. B.H., N.M. and S.J. optimized dissection and dissociation protocols. B.H. developed the experimental approach for combined scar and transcriptome detection, and B.H. and N.M. performed experiments. B.S. developed computational methods and analyzed the data, with support by P.O.-C. J.P.J. and N.N. guided experiments, and J.P.J. guided analysis. J.P.J. and B.S. wrote the manuscript, with input from all other authors. All authors discussed and interpreted results.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Jan Philipp Junker.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–20, Supplementary Table 1 and Supplementary Note 1

  2. 2.

    Life Sciences Reporting Summary

Excel files

  1. 1.

    Supplementary Dataset 1

    Technical information for all sequenced libraries

  2. 2.

    Supplementary Dataset 2

    Cell type information for 5 dpf larvae

  3. 3.

    Supplementary Dataset 3

    Scar detection statistics for 5 dpf larvae

  4. 4.

    Supplementary Dataset 4

    Cell type information for adult organs

  5. 5.

    Supplementary Dataset 5

    Scar detection statistics for adult organs

  6. 6.

    Supplementary Dataset 6

    Statistics connection enrichment analysis

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DOI

https://doi.org/10.1038/nbt.4124

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