Paired Single-Cell Transcriptome and DNA Barcode Detection in Zebrafish Using ScarTrace.

Area of Science:

• Developmental Biology
• Genetics
• Molecular Biology

Background:

• Understanding cell fate decisions during embryogenesis is crucial for developmental biology.
• Existing lineage tracing methods have limitations in resolution and scalability.

Purpose of the Study:

• To introduce ScarTrace, a novel CRISPR/Cas9-based genetic lineage tracing method.
• To demonstrate the utility of ScarTrace for reconstructing cell lineages and tissue development in zebrafish.

Main Methods:

• CRISPR/Cas9-mediated DNA barcoding of single cells in developing zebrafish embryos.
• Isolation of single cells from various adult zebrafish tissues (marrow, brain, eyes, fins).
• Single-cell cDNA amplification for transcriptome analysis and genomic DNA nested PCR for barcode sequencing.

Main Results:

• Successful unique DNA barcoding of single cells using a target GFP sequence.
• Capture of both transcriptome and barcode sequences from isolated adult cells.
• Computational identification of cell types and barcodes for clone tracing and lineage reconstruction.

Conclusions:

• ScarTrace provides a powerful tool for high-resolution genetic lineage tracing in zebrafish.
• The method allows for unraveling complex fate decisions during embryogenesis.
• ScarTrace facilitates the reconstruction of tissue lineage trees and developmental processes.