{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Pol Sorigue"],"organism":["Danio rerio"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-16780"],"description":["Using Oxford Nanopore Sequencing, we sequenced the methylome of wild type zebrafish (TL background) forebrain. Forebrains from six individuals were profiled, enabling direct detection of multiple DNA base modifications at single-base resolution. The resulting dataset includes CpG-associated 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), as well as non-CpG 5mC and N6-methyladenine (6mA). NOTE: Raw Oxford Nanopore sequencing reads have been deposited in the European Nucleotide Archive under accession PRJEB108899 - this record was created independently by the submitter, in case of access issues, please contact the submitter directly."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Library Construction - Sequencing libraries were prepared using the Rapid Barcoding Kit 24 v14 (SQK-RBK114-24; Oxford Nanopore Technologies) according to the manufacturer’s instructions. Each sequencing run included six individuals, resulting in 13 independent library preparation and sequencing runs. Sequencing was performed on a PromethION platform using FLO-PRO114M flow cells and MinKNOW software (version 25.05.14).","Growth Protocol - All animals used in this experiment were wild-type zebrafish (TL background) bred and maintained at the Instituto Gulbenkian de Ciência (IGC, Oeiras, Portugal). Fish were placed in a Tecniplast recirculating life support system at 5 days post-fertilization (dpf) and maintained in a polyculture of rotifers and algae until 8 dpf, after which the feeding regime was initiated. They were fed twice daily: rotifers in the morning and commercial food flakes (Gemma) in the afternoon. All animals were kept at 28 °C under a 14:10 h light-dark photoperiod, with a pH of 7.0 and conductivity of 1200 μS/cm. All experiments were conducted in accordance with the standard operating procedures of the Instituto Gulbenkian de Ciência Ethics Committee. No permit number was required for this experiment, as the animals were maintained under standard housing conditions and no experimental procedures were performed.","Sample Collection - A total of six fish (2 females and 4 males) were randomly selected from three different wild-type tanks. Fish were euthanized with an overdose of tricaine solution (MS-222, Pharmaq; 1200 mg/L), and the forebrain (telencephalon and diencephalon) was dissected under a stereomicroscope (Nikon, SMZ 745). After dissection, the forebrain tissue was collected and placed in 1.5 mL tubes containing 100 µL of HMW gDNA Tissue Lysis Buffer from the Monarch High Molecular Weight (HMW) DNA Extraction Kit for Tissue (New England Biolabs; NEB #T3060). Samples were immediately placed on dry ice and stored at −80 °C until further use. Biometric data were also collected, including standard length (measured from the tip of the snout to the end of the caudal peduncle) 2.53 ± 0.11, and body weight (g) 0.30 ± 0.02.","Nucleic Acid Extraction - Genomic DNA was extracted using the Monarch® High Molecular Weight (HMW) DNA Extraction Kit from Tissue (New England Biolabs, NEB #T3060), following the adapted manufacturer’s instructions to accommodate very low tissue inputs (< 2–5 ng of starting material). Briefly, brain tissues were mechanically dissociated using a sterile pestle. The kit employs a glass bead–based purification method optimized for the isolation of extremely high–molecular–weight DNA, combining gentle cell lysis with controlled DNA fragmentation and subsequent precipitation of DNA onto the surface of large glass beads. Protocol modifications were implemented to minimize mechanical agitation and preserve DNA integrity. Under these conditions, extracted DNA fragment sizes typically ranged from 50–250 kb using the standard protocol, with the potential to reach megabase-scale fragments when the lowest agitation speeds were employed. Purified DNA was eluted into low-binding tubes (Eppendorf) and stored at 4 °C until library preparation.","Sequencing - Sequencing libraries were prepared using the Rapid Barcoding Kit 24 v14 (SQK-RBK114-24; Oxford Nanopore Technologies) according to the manufacturer’s instructions. Each sequencing run included six individuals, resulting in 13 independent library preparation and sequencing runs. Sequencing was performed on a PromethION platform using FLO-PRO114M flow cells and MinKNOW software (version 25.05.14)."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Data Transformation - Basecalled reads were aligned to the zebrafish reference genome assembly GRCz12tu using Dorado with default alignment parameters. Only high-confidence primary alignments were retained by removing reads with mapping quality (MAPQ) < 10, secondary and supplementary alignments, unmapped reads, and reads shorter than 200 bp. Filtered BAM files were subsequently sorted and indexed using samtools (version 1.21). The modBAM files were converted to bedMethyl format using modkit with the pileup command. The bedMethyl format enables representation of per-base methylation levels by reporting genomic coordinates, strand information, read coverage, and modification frequencies. A confidence threshold of 0.8 was applied uniformly to call the presence or absence of a modification at each base. Three bedMethyl datasets were generated: 1. A comprehensive dataset containing all detected base modifications with strand-specific information. 2. A CpG-restricted dataset in which calls were limited to CpG dinucleotides and aggregated across strands. 3. A CH-restricted dataset (CA, CC, CT motifs) retaining strand-specific information. Strand aggregation for CpG sites was performed because CpG methylation is typically symmetric across complementary strands, allowing reads from both strands to contribute to a single genomic coordinate. The mitochondrial chromosome was excluded from all downstream analyses. For downstream processing, bedMethyl tables were reduced to include genomic position (0-based coordinate of the modified cytosine), read coverage, methylation fraction, strand, and motif context. The processed files available contain methylation matrices in BED format. Columns: 1: chromosome; 2: start (0-based coordinate), 3: end, 4: total reads (coverage), 5: methylation calls at that position, 6: strand (when applicable)."],"omics_type":["Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["PromethION"],"study_type":["methylation profiling by high throughput sequencing"],"species":["Danio rerio"],"pubmed_title":["Genome-wide DNA methylation profiling of the zebrafish forebrain"],"pubmed_authors":["Sorigue P., Pinget M., Costa JP, Teles MC, Oliveira RF.","Pol Sorigue"],"additional_accession":[]},"is_claimable":false,"name":"Zebrafish Forebrain Methylome","description":"Using Oxford Nanopore Sequencing, we sequenced the methylome of wild type zebrafish (TL background) forebrain. Forebrains from six individuals were profiled, enabling direct detection of multiple DNA base modifications at single-base resolution. The resulting dataset includes CpG-associated 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), as well as non-CpG 5mC and N6-methyladenine (6mA). NOTE: Raw Oxford Nanopore sequencing reads have been deposited in the European Nucleotide Archive under accession PRJEB108899 - this record was created independently by the submitter, in case of access issues, please contact the submitter directly.","dates":{"release":"2026-03-26T00:00:00Z","modification":"2026-03-26T02:03:18.554Z","creation":"2026-03-12T13:54:16.151Z"},"accession":"E-MTAB-16780","cross_references":{"ENA":["ERP189737"],"EFO":["EFO_0002944","EFO_0004170","EFO_0003789","EFO_0002761","EFO_0005518","EFO_0003816","EFO_0004184"]}}