Protocol to profile snATAC-seq datasets and motif enrichment analysis during zebrafish early embryogenesis

• Published in: STAR protocols 2024-12, Vol.5 (4), p.103501, Article 103501
• Main Authors: Zhou, Jie, Yang, Xueqian, Lin, Xiumei, Zhao, Kaichen, Wang, Xue, Dong, Zhiqiang, Liu, Chuanyu, Liu, Chang
• Format: Article
• Language: English
• Subjects: bioinformatics; developmental biology; model organisms; single cell
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2024.103501

The scarcity of zebrafish-specific motif databases presents a challenge to the analysis of transcription factor (TF) motif within zebrafish single-cell assay for transposase-accessible chromatin using sequencing (scATAC-seq) data, thus hindering the identification of regulatory elements throughout zebrafish embryonic development. Here, we provide a protocol to analyze single-nucleus chromatin accessibility dataset during zebrafish early embryogenesis. We describe steps for fragment file retrieval, sample integration, quality control, Latent Semantic Indexing (LSI) clustering, and peak calling via ArchR. Crucially, we detail procedures for zebrafish-specific motif database construction, motif enrichment, and TF footprinting analysis. For complete details on the use and execution of this protocol, please refer to Lin et al.1 [Display omitted] •Instructions for ArchR package installation and dataset preparation•Procedures to obtain a high-quality snATAC-seq dataset by quality control•Steps for dimensionality reduction, unsupervised clustering, and cell annotation•Codes for zebrafish-specific motif database construction and motif enrichment Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. The scarcity of zebrafish-specific motif databases presents a challenge to the analysis of transcription factor (TF) motif within zebrafish single-cell assay for transposase-accessible chromatin using sequencing (scATAC-seq) data, thus hindering the identification of regulatory elements throughout zebrafish embryonic development. Here, we provide a protocol to analyze single-nucleus chromatin accessibility dataset during zebrafish early embryogenesis. We describe steps for fragment file retrieval, sample integration, quality control, Latent Semantic Indexing (LSI) clustering, and peak calling via ArchR. Crucially, we detail procedures for zebrafish-specific motif database construction, motif enrichment, and TF footprinting analysis.