Thirty years after the first transgenic mouse was produced, a plethora of genetic tools has been developed to study immune cells in vivo. A powerful development is the bacterial artificial chromosome (BAC) transgenic approach, combining advantages of both conventional transgenic and knock-in gene-targeting strategies. In immunology the potential of BAC transgenic technology has yet to be fully harvested and, combined with a variety of elegant genetic tools, it will allow the analysis of complex immunological processes in vivo. In this short review, we discuss the applications of BACs in immunology, such as identification of regulatory regions, expression and cell-fate mapping, cell ablation, conditional mutations and the generation of humanized mice.