Using the Fly-FUCCI System for the Live Analysis of Cell Cycle Dynamics in Cultured Drosophila Cells

Cultured Drosophila cells are an attractive system for live imaging experiments, as this cell type is not very demanding in terms of temperature and media composition. Moreover, cultured Drosophila cell lines are very responsive to RNAi without being prone to off-target effects, and thus have become important for use in high-content screening. We have developed a fly-specific fluorescent, ubiquitination-based cell cycle indicator (FUCCI) system that enables faithful detection of G1, S, and G2 phases, and is thus a powerful tool for the analysis of cell cycle dynamics in living or fixed cells. Here, we describe a protocol for the generation of cell lines stably expressing the Fly-FUCCI sensors, followed by a description of how these cell lines can be employed in studies of cell cycle oscillation using live microscopy.