PUBLICATION
Gut Isolation from Zebrafish Larvae for Single-cell RNA Sequencing
- Authors
- Kakiailatu, N.J.M., Kuil, L.E., Bindels, E., Zink, J.T.M., Vermeulen, M., Melotte, V., Alves, M.M.
- ID
- ZDB-PUB-231128-2
- Date
- 2023
- Source
- Journal of visualized experiments : JoVE (201): (Journal)
- Registered Authors
- Alves, Maria, Kakiailatu, Naomi, Kuil, Laura
- Keywords
- none
- MeSH Terms
-
- Animals
- Gastrointestinal Tract*/physiology
- Intestines
- Larva/genetics
- Sequence Analysis, RNA
- Zebrafish*/genetics
- PubMed
- 38009742 Full text @ J. Vis. Exp.
Citation
Kakiailatu, N.J.M., Kuil, L.E., Bindels, E., Zink, J.T.M., Vermeulen, M., Melotte, V., Alves, M.M. (2023) Gut Isolation from Zebrafish Larvae for Single-cell RNA Sequencing. Journal of visualized experiments : JoVE. (201):.
Abstract
The gastrointestinal (GI) tract performs a range of functions essential for life. Congenital defects affecting its development can lead to enteric neuromuscular disorders, highlighting the importance to understand the molecular mechanisms underlying GI development and dysfunction. In this study, we present a method for gut isolation from zebrafish larvae at 5 days post fertilization to obtain live, viable cells which can be used for single-cell RNA sequencing (scRNA-seq) analysis. This protocol is based on the manual dissection of the zebrafish intestine, followed by enzymatic dissociation with papain. Subsequently, cells are submitted to fluorescence-activated cell sorting, and viable cells are collected for scRNA-seq. With this method, we were able to successfully identify different intestinal cell types, including epithelial, stromal, blood, muscle, and immune cells, as well as enteric neurons and glia. Therefore, we consider it to be a valuable resource for studying the composition of the GI tract in health and disease, using the zebrafish.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping