{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Dr. Steve Ayobahan"],"organism":["Danio rerio"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-15674"],"description":["GenX (HFPO-DA) is a per- and polyfluoroalkyl substance (PFAS) increasingly used as a replacement for legacy compounds such as PFOA, but its environmental persistence and potential toxicity raise concerns. Traditional chemical risk assessment is time- and resource-intensive and provides limited insight into underlying mechanisms. In this study, zebrafish embryos were exposed to sublethal concentrations of GenX for 96 hours, and RNA sequencing was used to assess concentration-dependent changes in gene expression. These data allow identification of sensitive transcriptional responses and affected biological pathways, supporting the use of transcriptomic approaches for mechanistic, 3R-compliant evaluation of GenX toxicity."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Library Construction - Adapter sequences were removed using trimmomatic (v0.39) and library’s sequence quality was assessed with FastQC (v0.12.0). Reads were checked for potential contaminations with FastQ Screen (v0.15.3) using bowtie2 (v2.5.3) against the following genomes: Daphnia magna (ftp://ftp.ensemblgenomes.org/pub/metazoa/release-48/fasta/daphnia_magna/dna/Daphnia_magna.daphmag2.4.dna.toplevel.fa.gz), Homo sapiens, Mus musculus, Drosophila melanogaster, Saccharomyces cerevisiae, Eschericia coli as well as custom build human rRNA database. With the exception of D. magna, pre-built Bowtie2 indices for those genomes were directly downloaded from Babraham Bioinformatics with the built-in function ‘fastq_screen --get_genomes’.","Sample Collection - 2,3,3,3-tetrafluoro-2-(heptafluoropropoxy)propanoic acid (HFPO-DA, also referred to as GenX, CAS 13252-13-6) was purchased from Sigma-Aldrich, St. Louis, MO, USA. The test solution was prepared by first preparing a highly concentrated stock solution in water, fresh one day prior to the start of each experiment. The test solutions were prepared from these stock solutions immediately before the start of the test to avoid any degradation of the substance. All GenX test solutions were prepared in 100 % methanol (Chromasolv®, Sigma-Aldrich, CAS 67-56-1). Using a binocular optical microscope, 15 embryos of the same early blastula stage were transferred from the pre-picking plate to the test Polystyrene plates filled with 8 ml of test solution. The embryos were then incubated as described in the “growth protocol”.","Growth Protocol - Adult animal husbandry: Wild-type zebrafish (Danio rerio, Strain AB) broodstocks were maintained under flow-through conditions in 150 L tanks at 26 +/- 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® (Tetra Werke, Melle, Germany) main feed ad libitum and Artemia salina nauplii. One day before the start of the test, glass spawning trays containing artificial substrate (green glass beads strung on stainless steel wire) were placed at the bottom of each tank. After mating and spawning in the morning hours eggs were rinsed with clear Cu-reduced water placed into the dishes for pre-picking.","Sequencing - Poly(A)+ RNA was purified and fragmented. It was then transcribed into cDNA using the TruSeq RNA library Prep Kit v2 from Illumina. After adapter ligation and PCR amplification, the RNA fragments were sequenced using an Illumina NovaSeq 6000 system. The sequencing was done in paired-end mode with a read length of 150 base pairs. A minimum read depth of 30,000,000 read pairs per sample was ensured. Adapter trimming and sequencing quality control were performed using fastp and FastQC, respectively. Contamination assessment was done using FastQScreen. The overall quality report was generated using MultiQC.","Nucleic Acid Extraction - Total RNA and Protein were extracted from the tissue lysate using the NucleoSpin RNA Protein kit from Macherey-Nagel following the manufacturer's protocol. The RNA concentration was measured using a Nanodrop 2000 spectrophotometer from Thermo Scientific, with a minimum concentration of 100 ng/µl required. The overall RNA quality and RIN value were determined using the Agilent 2100 Bioanalyzer system and the Agilent RNA 6000 Nano Kit, respectively. Only samples with a RIN value greater than 8 were used for downstream analysis. The samples were stored at -80°C until they were sent to the sequencing facility on dry ice. For each sample, 10 zebrafish larvae were collected and transferred to a low binding Eppendorf tube. The larvae were euthanized simultaneously by placing the tubes on ice for 10 minutes. The supernatant was carefully removed without injuring the larvae, and RP1 buffer from the NucleoSpin RNA Protein kit (Macherey-Nagel 740933.250) was added for tissue homogenization. The RP1 buffer was prepared with TCEP as a reducing agent. Detailed information about embryonic incubation conditions, husbandry of adult broodstocks, exposure duration, and conditions can be found in the \\\"Growth protocol\\\" and \\\"Treatment protocol\\\" respectively.","Sample Treatment - Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes (diameter 6 cm) filled with 8 ml of the respective nominal concentrations of 2,5 µg/L, 25 µg/L, 250 µg/L, 2500 µg/L, and 25000 µg/L. Embryos were incubated at 27 ± 1°C on a 14:10 h light/dark cycle. At 24 hours post fertilization (hpf), eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf, overall survival, hatching rates, morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh, aerated test solutions. At 96 hpf again, overall survival, hatching rates, morphological malformations and physiological changes were recorded before RNA extraction."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Data Transformation - Gene level raw counts from the experiment’s libraries were bind into a single count matrix in R (version 4.2.2) where rows correspond to genes and columns to the sample (GenX_CountMatrix.txt). Only genes with a minimum sum of N counts across N samples (rowSums >= N) were kept for further downstream analysis. After low read counts were removed, the remaining feature mapped gene counts in the matrix were normalized with the DESeq2 package (1.42.1) using a parametric fit type model for dispersion estimates (default). The resulting normalized count matrix (GenX _DESeqNormCounts.txt) was used for differential gene expression analysis with DESeq2 and further downstream data analysis.","Sequence Alignment - Sequences were aligned to the reference genome Danio rerio GRCz11 (version 91, https://www.ensembl.org/Danio_rerio/Info/Index) using STAR aligner (v2.7.11) allowing for 2 mismatches. Subsequently, feature mapped reads were directly counted by STAR’s ‘-- quantMode GeneCounts’ function which produces counts coincide with those produced by HTSeq’s ‘htseq-count’ with default parameters (union mode)."],"omics_type":["Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["Illumina NovaSeq 6000"],"study_type":["RNA-seq of coding RNA"],"species":["Danio rerio"],"pubmed_authors":["Dr. Sebastian Eilebrecht","Dr. Steve Ayobahan"],"additional_accession":[]},"is_claimable":false,"name":"Transcriptomic responses of zebrafish embryos to sublethal exposure to GenX (HFPO-DA): A 96-Hour RNA-seq Study","description":"GenX (HFPO-DA) is a per- and polyfluoroalkyl substance (PFAS) increasingly used as a replacement for legacy compounds such as PFOA, but its environmental persistence and potential toxicity raise concerns. Traditional chemical risk assessment is time- and resource-intensive and provides limited insight into underlying mechanisms. In this study, zebrafish embryos were exposed to sublethal concentrations of GenX for 96 hours, and RNA sequencing was used to assess concentration-dependent changes in gene expression. These data allow identification of sensitive transcriptional responses and affected biological pathways, supporting the use of transcriptomic approaches for mechanistic, 3R-compliant evaluation of GenX toxicity.","dates":{"release":"2026-05-01T00:00:00Z","modification":"2026-05-01T01:03:24.257Z","creation":"2025-10-08T14:49:45.863Z"},"accession":"E-MTAB-15674","cross_references":{"ENA":["ERP181221"],"EFO":["EFO_0002944","EFO_0004170","EFO_0003789","EFO_0004917","EFO_0005518","EFO_0003816","EFO_0003738","EFO_0004184","EFO_0003969"]}}