Revolution spinning disc system

The 96-h AEL larval brains expressing one to three copies of survivin::mDendra2 were used. Photoconversion experiments were performed on an Andor Revolution spinning disc system containing the FRAPPA unit (Andor). Several regions of interests (ROIs) were manually chosen in the GFP channel. Survivin at the metaphase plate was irradiated before anaphase onset. Before photoconversion, single Z planes containing ROIs were scanned for ten time points with maximum speed. Subsequently, ROIs were irradiated with the 405 nm laser line (9.7%; 20 repeats; 50 μs dwell time). After photoconversion, the entire neuroblast was scanned with a z-step size of 0.65 μm. Photoconverted Survivin-mDendra2 emits red fluorescence. Converted and unconverted mDendra2 emission were merged in Andor IQ2 and converted into Imaris.

Ninety-six hours larval brains expressing Zipper::mDendra2 were used after their dissociation (see above). The photoconversion experiments were performed on an Andor Revolution spinning disc system containing Andor’s FRAPPA unit. Several regions of interests (ROIs) were manually chosen in the GFP channel and Zipper::mDendra2 was irradiated with 405 nm on either the apical, basal or lateral cortex just after anaphase onset. Before photoconversion, single Z planes containing ROIs were scanned for ten time points with maximum speed. Subsequently, ROIs were irradiated with 405 nm (10%; 50 repeats; 50 µs dwell time). After photoconversion, the entire neuroblast was scanned with a z-step size of 0.65 µm. Converted and unconverted mDendra2 emission were merged in AndorIQ2 and converted into Imaris.