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

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.