Functional studies of resident microglia require molecular tools for their genetic manipulation. Here we show that microRNA-9-regulated lentiviral vectors can be used for the targeted genetic modification of resident microglia in the rodent brain. Using transgenic reporter mice, we demonstrate that murine microglia lack microRNA-9 activity, whereas most other cells in the brain express microRNA-9. Injection of microRNA-9-regulated vectors into the adult rat brain induces transgene expression specifically in cells with morphological features typical of ramified microglia. The majority of transgene-expressing cells colabels with the microglia marker Iba1. We use this approach to visualize and isolate activated resident microglia without affecting circulating and infiltrating monocytes or macrophages in an excitotoxic lesion model in rat striatum. The microRNA-9-regulated vectors described here are a straightforward and powerful tool that facilitates functional studies of resident microglia.