RNA-Seq of zebrafish embryos (96hpf) treated with different concentrations of Triiodothyronine (T3) below acute toxicity levels against untreated control groups. RNA-Seq of zebrafish embryos (96hpf) treated with different concentrations of Triiodothyronine (T3) below acute toxicity levels against untreated control groups

The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish (Danio rerio) embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine (T3, CAS: 6893-02-3) was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test (OECD 236), 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for 96 hours under semi-static conditions. Each test comprised of a low exposure (LE), high exposure (HE) and negative control (NC) group and was performed in triplicates. At 96 hours post fertilization (hpf), 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit (Macherey-Nagel). RNA quality was assessed with a 2100 Bioanalyzer system (Agilent) before coding RNA was purified (PolyA selection with TruSeq RNA Library Prep Kit v2) and sequenced on an Illumina HiSeq 4000 System (Illumina) in 50 bp single read mode, producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.