Generation of transgenic mice expressing FRET biosensors.

Transgenic mice play a significant role in modern biomedical research. They allow not only mechanistic insights into the functions of specific genes and proteins. Recent strategies have also established the use of transgenic mice as an exciting tool for the expression and in vivo or in situ analysis of fluorescent biosensors, which are capable of directly reporting second messenger levels and biochemical processes in real time and in living cells. In this chapter, we present a detailed protocol for the generation of plasmid vectors and transgenic mice expressing a Förster resonance energy transfer (FRET)-based biosensor for the second messenger 3',5'-cyclic adenosine monophosphate. These tools and techniques should provide great potential for the analysis of second messenger dynamics in a more physiologically relevant context.