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Single-Cell Lineage Tracing and Clonal State-Fate Analysis.

Alejo E Rodriguez Fraticelli1,2, Pedro Sánchez Sánchez3

  • 1Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology, Barcelona, Catalonia, Spain. alejo.rodriguez-fraticelli@irbbarcelona.org.

Methods in Molecular Biology (Clifton, N.J.)
|January 2, 2025
PubMed
Summary

Clonal lineage tracing using random barcodes enables tracking of numerous cell clones over time. This method, combined with single-cell omics, reveals cell states and fate determinants for a comprehensive understanding of biological processes.

Keywords:
BarcodeClonal memoriesLineage tracingSequencingSingle cellState-fateTranscriptomeClones

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Area of Science:

  • Molecular Biology
  • Genetics
  • Developmental Biology
  • Immunology

Background:

  • Lineage tracing is crucial for understanding cellular processes like development and immunity.
  • Traditional methods require prior cell type knowledge and limit the number of trackable clones.
  • Clonal lineage tracing with random barcodes offers a scalable alternative for tracking cell populations.

Purpose of the Study:

  • To provide a roadmap for designing and implementing single-cell lineage tracing experiments.
  • To highlight key parameters for stable barcode integration, clone expansion, and omics data capture.
  • To guide researchers in analyzing sequencing data for clonal identity assignment and fate determinant identification.

Main Methods:

  • Utilizing lentiviral libraries for stable integration of expressed random barcodes.
  • Combining clonal barcoding with single-cell omics technologies (e.g., transcriptomics).
  • Optimizing experimental design for cellular sampling, barcode diversity, and minimizing clonal dropout.

Main Results:

  • Demonstration of clonal lineage tracing's capability to label and track a vast array of cells and their progeny.
  • Presentation of available lentiviral barcode libraries suitable for capturing transcriptome data.
  • Discussion of computational tools and tips for sequencing data analysis, including clonal assignment and fate determinant identification.

Conclusions:

  • Clonal lineage tracing with random barcodes is a powerful alternative to traditional methods, enabling high-throughput cell tracking.
  • Integration with single-cell omics provides deep insights into cell states and their maintenance over time.
  • The study offers practical guidance for experimental design and data analysis, facilitating comprehensive exploration of fate determinants.