Performing Chromophore-Assisted Laser Inactivation in Drosophila Embryos Using GFP.
Methods in molecular biology, January 2016
Anne Pélissier-Monier, Bénédicte Sanson, Bruno Monier, Pélissier-Monier, Anne, Sanson, Bénédicte, Monier, Bruno
Chromophore-assisted laser inactivation (CALI) is an optogenetic technique in which light-induced release of reactive oxygen species triggers acute inactivation of a protein of interest, with high spatial and temporal resolution. At its simplest, selective protein inactivation can be achieved via the genetic fusion of the protein to a photosensitizer such as EGFP, and using standard optical setups such as laser scanning confocal microscopes. Although use of CALI in Drosophila is relatively recent, this technique can be a powerful complement to developmental genetics, especially in vivo as it allows visualization of the immediate consequences of local protein inactivation when coupled to time-lapse microscopy analysis. In addition to providing examples of protocols, this chapter is intended as a conceptual framework to support the rational design of CALI experiments.
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