Kaede-centrin1 labeling of mother and daughter centrosomes in mammalian neocortical neural progenitors.

The importance of the centrosome in regulating basic cellular processes and cell fate decisions has become increasingly evident from recent studies tracing the etiology of developmental disorders to mutations in genes encoding centrosomal proteins. This unit details a protocol for a fluorescence-based pulse labeling of centrioles of neural progenitor cells in the developing neocortex of mice. In utero electroporation of Kaede-Centrin1 followed by in utero or ex vivo photoconversion allows a direct monitoring of the inheritance of centrosomes containing centrioles of different ages in dividing neocortical neural progenitors (i.e., radial glial cells). This is achieved by combining the irreversible photoconversion capacity of the Kaede protein from green to red fluorescence with the faithful duplication of the centrosome during each cell cycle. After two mitotic divisions following photoconversion, mother centrosomes containing the original labeled centriole appear in both red and green fluorescence, and can be distinguished from daughter centrosomes which appear in green fluorescence only. This facilitates the study of the inheritance and behavior of the mother and daughter centrosomes in asymmetric cell divisions in the developing mammalian neocortex.