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Related Experiment Video

Updated: Jun 14, 2025

Sample Preparation and Analysis of RNASeq-based Gene Expression Data from Zebrafish
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Isolation and Preparation of Embryonic Zebrafish Retinal Cells for Single-Cell RNA Sequencing.

Shea A Heilman1, Dennis Kostka2,3, Hannah Schriever3

  • 1Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|September 6, 2024
PubMed
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This study presents a reproducible protocol for generating high-quality single-cell suspensions from embryonic zebrafish eyes. This method ensures viable cells for single-cell RNA sequencing (scRNA-Seq), enabling robust biological discovery.

Area of Science:

  • Developmental Biology
  • Genomics
  • Zebrafish Research

Background:

  • Single-cell RNA sequencing (scRNA-Seq) offers high precision for biological questions.
  • Ocular development and regeneration research has benefited significantly from scRNA-Seq discoveries.
  • A key challenge in scRNA-Seq is obtaining high-quality, viable cell populations for analysis.

Purpose of the Study:

  • To describe a reproducible methodology for generating high-quality single-cell suspensions from embryonic zebrafish eyes.
  • To provide a protocol that addresses the uncertainty and technical hurdles associated with scRNA-Seq experiments.
  • To enable rigorous computational analyses by ensuring usable data from scRNA-Seq.

Main Methods:

  • Detailed protocol for dissociating embryonic zebrafish eyes into single-cell suspensions.
Keywords:
Cell viabilityRetinaZebrafishscRNA-seq

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  • Utilized the 10× Genomics v3.1 system for scRNA-Seq.
  • Established quantitative quality control parameters for cell yield and viability.

    • Main Results:

    • Successfully generated high-quality single-cell suspensions from embryonic zebrafish eyes.
    • Proof-of-principle studies demonstrated high-quality scRNA-Seq data output.
    • Methodology allows for quantitative assessment of critical quality control parameters.

    Conclusions:

    • The described protocol provides a reliable starting point for scRNA-Seq experiments in zebrafish eye research.
    • This methodology can be adapted for scRNA-Seq in other developing or regenerating tissues in zebrafish and other model systems.
    • Facilitates reproducible and high-quality scRNA-Seq data generation for advancing biological insights.