In vitro studies have previously shown that transcription factors find their binding sites through a mechanism of 'facilitated' diffusion that combines three-dimensional diffusion with sliding along DNA. Using a single-molecule approach, Hammar et al. have now shown that this mechanism also occurs in living bacterial cells. They used a fluoroescently labelled lac repressor protein (LacI) and monitored its binding to the lac operator. They show that LacI slides on average 45 ± 10 bp, and in most cases it slides over the operator sequence at least once before binding to it. During evolution, transcription factors may need to maintain the ability to slide over non-specific sequences as well as to bind specifically.