Bacteriophage Mu exhibits low specificity for the 5 bp sequence it selects as its transposition target, but shows regional biases in its insertion choices. For example, Mu prefers AT-rich DNA in vitro, exhibits a 1000-fold bias in target preference within the E. coli chromosome, and avoids targets carrying Mu end sequences. The Mu transposase is responsible for recognition of the 5 bp target consensus, but depends on the accessory protein MuB for efficient target capture. MuB preferentially binds to AT-rich DNA, explaining this particular regional preference. We have uncovered opposing roles for MuB in target capture and integration. We show while MuB-bound AT-rich DNA is favored for integration, the bound DNA itself is refractory, and that transposition occurs adjacent to, but not within the bound region. We show that this property of MuB is likely responsible for immunity of Mu from self-integration, since MuB was found to be strongly bound within the Mu genome. Genome-wide analysis of MuB binding on the E. coli chromosome showed that Mu target preference is positively related to MuB binding profile, and that MuB binding is insulated by the nucleoid-associated protein Fis but not by transcription events. Since Fis binding to the chromosome responds to the frequency of A-tracts, a chromosome domain structure signal, Mu transposition must also respond to chromosome domain signals. Work in this dissertation has provided a new understanding of how MuB influences and controls Mu target choice, and of reciprocal interactions between a bacterial chromosome and a transposable element. / text