biostudies-arrayexpressAdelino CanarioDanio reriohttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-15114The objective was to identify downstream gene targets of pSmad1/5 and pStat3 using the CUT & TAG techniquebiostudies-arrayexpressLibrary Construction - The nucleic library was constructed using primers and reagents provided with the NovoNGS® CUT&Tag® 2.0 High-Sensitivity Kit (for Illumina®）from Novoprotein Scientific Inc., Shanghai, China. Take out the cryovial and shake it in 37°C water bath to quickly thaw it within 1-2min.Cells were centrifuged for 5 min at 600×g at room temperature. Cell activity was detected with LUNA-FLTM and counted. CUT&Tag assay was performed as described previously(Kaya-Okur, Wu et al. 2019). Briefly, the cells are bound to Concanavalin A-coated magnetic beads, and the cell membrane is permeabilised by Digitonin. The enzyme pA-Tn5 Transposase precisely binds the DNA sequence near the target protein under the antibody guidance and results in factor-targeted tagmentation. DNA sequence is taNucleic Acid Extraction - DNA was purified using Tagment DNA extraction beads provided in the NovoNGS® CUT&Tag® 2.0 High-Sensitivity Kit (for Illumina®）from Novoprotein Scientific Inc., Shanghai, China.Sequencing - The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. The library preparations were sequenced on Illumina Novaseq platform at Novogene Science and Technology Co., Ltd (Beijing, China) and 150 bp paired-end reads were generatedSample Collection - Zebrafish embryos of the AB strain were collected and the yolk was removed. Cells were dissociated into single cells with trypsin and collagenase.Dissociated single cells were combined with CoA beads and incubated with primary antibody anti-pStat3 (HUABIO, 1:50) or anti-pSmad1/5 (Sigma-Aldrich, 1:50) overnight. Samples were incubated with secondary antibody Goat anti-rabbit (Bio-Rad, 1:100), conjugated with transposase, and fragmented.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - Quality control Raw data (raw reads) of fastq format were firstly processed using fastp (version 0.20.0). In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low-quality reads from raw data. At the same time, Q20, Q30 and GC content of the clean data were calculated. All the downstream analyses were based on the clean data. Reads mapping to the reference genome Reference genome and gene annotation files were downloaded from the zebrafish genome website directly. Index of the reference genome was built using BWA (v0.7.12) and clean reads were aligned to the reference genome using BWA mem. These reads were then filtered for high quality (MAPQ ≥ 13),we also removed reads that were not properly paired andMetabolomicsUnknownTranscriptomicsGenomicsProteomicsIllumina NovaSeq 6000RNA-seq of coding RNADanio rerioJie ChenAdelino CanariofalseCUT&TAG of zebrafish pSmad1/5 and pStat3The objective was to identify downstream gene targets of pSmad1/5 and pStat3 using the CUT & TAG technique2026-07-22T00:00:00Z2026-07-22T22:37:16.484Z2025-05-02T13:36:33.248ZE-MTAB-15114ERP172213E-MTAB-15099EFO_0002944EFO_0004170EFO_0005518EFO_0003816EFO_0003738EFO_0004184