Related Experiment Video
Updated: Jun 28, 2025
09:38
Fluorescence Labeling to Visualize Low-Expressed Proteins in Zebrafish
Published on: January 24, 2025
546
Tagging the tjp1a Gene in Zebrafish with Monomeric Red Fluorescent Protein Using Biotin Homology Arms
Connor Davison1, Hamelynn Harzman1, Jessie Nicholson1
1Department of Genetics, Development and Cell Biology, Iowa State University, Ames, Iowa, USA.
Zebrafish
|April 15, 2024
Summary
Researchers developed a new method to fluorescently label Tjp1a proteins in zebrafish embryos. This technique enables visualization of cell adhesion dynamics during development, aiding studies on vascular formation.
Area of Science:
- Cell Biology
- Developmental Biology
- Genetics
Background:
- Tight junction proteins like Tjp1a are crucial for maintaining epithelial and endothelial tissue integrity.
- Understanding the dynamic behavior of these proteins is essential for studying tissue development and disease.
Purpose of the Study:
- To develop a method for fluorescently labeling Tjp1a in zebrafish.
- To enable real-time observation of cell-cell adhesion dynamics during embryonic development.
Main Methods:
- CRISPR/Cas9-mediated targeted integration was used to insert monomeric red fluorescent protein (mRFP) into the zebrafish *tjp1a* gene.
- Biotin-labeled PCR-generated templates with short homology arms were employed for precise genomic integration.
- Microinjection of Cas9-gRNA complexes and repair templates into one-cell zebrafish embryos.
Main Results:
- A precise integration allele of *tjp1a-mRFP* was successfully generated and recovered.
- Confocal imaging confirmed RFP labeling at cell-cell junctions, visualizing Tjp1a localization.
- The established *tjp1a-mRFP* zebrafish line allows for tracking junctional dynamics.
Conclusions:
- The developed CRISPR/Cas9 method provides a reliable way to fluorescently tag Tjp1a in zebrafish.
- This tool facilitates the study of cell adhesion and tissue morphogenesis, particularly in vascular development.
