High-throughput sequencing technologies [next-generation sequencing (NGS)] are increasingly used to address diverse biological questions. Despite the rich information in NGS data, particularly with the growing datasets from repositories like the Genome Sequence Archive (GSA) at NGDC, programmatic access to public sequencing data and metadata remains limited. We developed iSeq to enable quick and straightforward retrieval of metadata and NGS data from multiple databases via the command-line interface. iSeq supports simultaneous retrieval from GSA, SRA, ENA, and DDBJ databases. It handles over 25 different accession formats, supports Aspera downloads, parallel downloads, multi-threaded processes, FASTQ file merging, and integrity verification, simplifying data acquisition and enhancing the capacity for reanalyzing NGS data. iSeq is freely available on Bioconda (https://anaconda.org/bioconda/iseq) and GitHub (https://github.com/BioOmics/iSeq).
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