The Rfam database, a comprehensive resource for non-coding RNA (ncRNA) families, has been updated to version 12.0. This release includes significant improvements and new features, such as an upgraded search pipeline using Infernal 1.1, which enhances homology detection and speeds up the annotation process. The new pipeline is more user-friendly and can handle large genomic and metagenomic datasets efficiently. Rfam 12.0 introduces 260 new families, including well-studied large subunit ribosomal RNA (LSU rRNA) and various bacterial and archaeal small RNA families. The database now also includes information on short sequence- and structure-based RNA motifs within families, a first for Rfam. The update has been accompanied by adjustments to curator-defined gathering thresholds to optimize homology detection. Additionally, Rfam has incorporated RNAcentral, a resource for ncRNA sequence data, to enhance annotation and facilitate cross-referencing. These enhancements make Rfam 12.0 a more powerful and user-friendly tool for ncRNA research.The Rfam database, a comprehensive resource for non-coding RNA (ncRNA) families, has been updated to version 12.0. This release includes significant improvements and new features, such as an upgraded search pipeline using Infernal 1.1, which enhances homology detection and speeds up the annotation process. The new pipeline is more user-friendly and can handle large genomic and metagenomic datasets efficiently. Rfam 12.0 introduces 260 new families, including well-studied large subunit ribosomal RNA (LSU rRNA) and various bacterial and archaeal small RNA families. The database now also includes information on short sequence- and structure-based RNA motifs within families, a first for Rfam. The update has been accompanied by adjustments to curator-defined gathering thresholds to optimize homology detection. Additionally, Rfam has incorporated RNAcentral, a resource for ncRNA sequence data, to enhance annotation and facilitate cross-referencing. These enhancements make Rfam 12.0 a more powerful and user-friendly tool for ncRNA research.