SAMtools and BCFtools are widely used software tools for processing and analyzing high-throughput sequencing data. They were first released twelve years ago and have since been continuously maintained and improved, incorporating numerous new features and enhancements. These tools are essential for tasks such as file format conversion, sorting, querying, statistics, variant calling, and effect analysis. Both packages are freely available under the MIT license and have been installed over a million times via Bioconda. They are written in C, optimized for low memory consumption and high speed, and have extensive test suites to ensure reliability. The development of these tools has been driven by the needs of large-scale sequencing projects and individual user requests, and they have been used to process and analyze sequencing data from various species. Future versions are expected to further enhance parallelization capabilities and address limitations in handling extremely large files and highly polymorphic sites. The source code and documentation for both SAMtools and BCFtools are available on GitHub and at <https://www.htslib.org>.SAMtools and BCFtools are widely used software tools for processing and analyzing high-throughput sequencing data. They were first released twelve years ago and have since been continuously maintained and improved, incorporating numerous new features and enhancements. These tools are essential for tasks such as file format conversion, sorting, querying, statistics, variant calling, and effect analysis. Both packages are freely available under the MIT license and have been installed over a million times via Bioconda. They are written in C, optimized for low memory consumption and high speed, and have extensive test suites to ensure reliability. The development of these tools has been driven by the needs of large-scale sequencing projects and individual user requests, and they have been used to process and analyze sequencing data from various species. Future versions are expected to further enhance parallelization capabilities and address limitations in handling extremely large files and highly polymorphic sites. The source code and documentation for both SAMtools and BCFtools are available on GitHub and at <https://www.htslib.org>.