An Improved, High-Yield Method for Isolating Nuclei from Individual Zebrafish Embryos for Single-Nucleus RNA Sequencing.

Abstract

Zebrafish is 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 single-cell combinatorial indexing RNAseq 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.