bustools

Bustools is a collection of open-source command-line tools for processing single-cell RNA sequencing data using the BUS format (barcodes, UMIs, and gene sets). It is designed to work with the BUS files produced by the kallisto bus workflow and to convert raw sequencing reads into gene–cell expression matrices. The BUS format is a compact representation of barcodes, UMIs, and transcript assignments for each read, enabling efficient counting at scale.

Bustools provides modular utilities for end-to-end processing of scRNA-seq data. Key components include tools for sorting BUS files, correcting barcode sequences against a reference whitelist, and counting molecules by gene for each cell. It supports UMI-aware counting and can handle multiple samples, allowing users to merge or separate results as needed. The suite is compatible with popular scRNA-seq platforms such as 10x Genomics and Drop-seq, and it can incorporate transcript-to-gene mappings to generate gene-level counts.

Typical workflow: sequencing reads are pseudoaligned by kallisto to produce a BUS file; bustools sorts and, if desired, corrects barcodes; bustools count tallies unique molecular identifiers per gene per barcode to produce a cell-by-gene count matrix, which can then be used in downstream analyses with standard tools. Bustools emphasizes efficiency and scalability, enabling processing of large datasets with modest computational resources.

Bustools is an open-source project hosted on GitHub as part of the kallisto-bustools ecosystem, maintained by contributors from the single-cell analysis community. It is widely referenced in single-cell RNA-seq pipelines for its speed and straightforward interface.