The ability to efficiently investigate transcription factor binding sites (TFBSs) genome-wide is central to computational studies of gene regulation. TFBSTools is an R/Bioconductor package for the analysis and manipulation of TFBSs and their associated transcription factor profile matrices. TFBStools provides a toolkit for handling TFBS profile matrices, scanning sequences and alignments including whole genomes, and querying the JASPAR database. The functionality of the package can be easily extended to include advanced statistical analysis, data visualization and data integration. AVAILABILITY AND IMPLEMENTATION: The package is implemented in R and available under GPL-2 license from the Bioconductor website (http://bioconductor.org/packages/TFBSTools/). CONTACT: ge.tan09@imperial.ac.uk SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.