An improved, high yield method for isolating nuclei from individual zebrafish embryos for single-nucleus RNA sequencing

TL;DR

This paper presents a new high-yield, efficient method for isolating nuclei from individual zebrafish embryos suitable for single-nucleus RNA sequencing, improving upon previous enzymatic dissociation techniques.

Contribution

The authors developed a bead homogenization-based protocol for isolating high-quality nuclei from zebrafish embryos in a 96-well format, compatible with various developmental stages.

Findings

Method yields high-quality nuclei efficiently

Compatible with multiple developmental stages

Outperforms enzymatic dissociation

Abstract

Zebrafish are an ideal system to study the effect(s) of chemical, genetic, and environmental perturbations on development due to their high fecundity and fast growth. Recently, single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high-quality nuclei from zebrafish embryos in a 96-well plate format by bead homogenization in a lysis buffer. Through head-to-head sciPlex-RNA-seq experiments, we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.