Overview
Single cell RNA-seq of wild type and perturbed zebrafish development
We explored the individual embryo and cell type-specific responses of zebrafish embryos to genetic and temperature perturbations using a scalable combinatorial-indexing scRNA-seq (sci-RNA-seq) approach. By barcoding hundreds-thousands of individual embryos, we statistically assess the changes in cell type abundances in response to each perturbation (3 temperatures, or 23 genetic loss of function experiments). Totalling ~3 million single cells across almost 2000 individual embryos, we put forward an analytical framework for high resolution phenotyping of diverse developmental traits in whole organisms.
Our findings are published in Saunders, Srivatsan, et al. Nature (2023) and Dorrity, et al. Cell (2023).
For code and processed tables, please visit our GitHub repositories: (1) Reference and genetic perturbations or (2) Temperature perturbations.
Study Design
Our studies produced three main datasets:
1. Reference atlas - A hierarchically annotated, individually-resolved reference atlas, comprising 1,167 individuals and 1.2 million cells over 19 timepoints (from 18-96 hpf).
2. Genetic perturbation atlas - Two million cells from fish recieving 23 different genetic perturbations over multiple timepoints, totaling 645 individually barcoded animals (we collected 8 or more embryos per condition/timepoint).
| Tissue | gene targets |
|---|---|
| Mesoderm | cdx4, cdx4;cdx1a, tbxta, tbx16, tbx16;tbx16l, tbx16;msgn1, wnt3a;wnt8a, noto, smo, tbx1, hand2 |
| Central or peripheral nervous system | egr2b, epha4a, hoxb1a, mafba, zc4h2, phox2a, foxi1, hgfa, met |
| Neural crest lineages | tfap2a, tfap2a;foxd3 |
3. Temperature perturbation atlas - 400,000 cells from fish raised at three different temperatures over three timepoints, totaling 288 individually barcoded animals.
Cell metadata breakdown
Our cell metadata contains lots of information about time, perturbation, statistical metrics, and annotation. Here is a breakdown of those attributes according to our column names:
timepoint: The developmental stage in hours post fertilization (hpf) of embryos. Embryos were staged according to key landmarks according to (Kimmel, et al (1995)). Options are 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 72, and 96 hpf.expt: Denotes unique library preparation instances.cell_type_sub: The finest cell type annotation level.cell_type_broad: A broader level of annotation than "cell_type_sub", but still capturing all uniquely identified cell types.tissue: The tissue type annotation that contains the cell types.germ_layer: The germ layer of origin for each cell type, if known.gene_target: The genetic perturbation target. Controls include injected (scrambled) and unjected. Sibling controls are listed as "ctrl-[target]", for null mutants included in the study.mean_nn_time: The mean time points of the nearest 15 neighbor cellsembryo: The individual embryo barcode.temp: The growth temperature of the embryos (standard conditions are 28C).pseudostage: Embryo-level staging prediction by cell composition.
Background art by Elena Bansh.