Ers spinning disk confocal system

Syncytial stage embryos were imaged on a Perkin Elmer ERS Spinning Disk confocal system (ERS software) mounted on a Zeiss Axiovert microscope, using a 63× 1.4 NA oil-immersion objective. Alternatively embryos were imaged on a Perkin Elmer Ultra-VIEW VoX (Volocity software) mounted on an IX81 microscope (Olympus), using a 60× 1.3 NA silicon immersion objective and an electron-multiplying charge-coupled device camera (ImagEM, Hamamatsu Photonics). All control/experiment pairs were analyzed on the same microscope system with identical illumination and acquisition settings.
For experiments to analyze the effects of PLP deletions, mRNAs encoding GFP-PLP deletions were injected into embryos and then imaged 60–120 min later.

Syncytial stage embryos were imaged on a Perkin Elmer ERS Spinning Disk confocal system (ERS software) mounted on a Zeiss Axiovert microscope, using a 63×, 1.4NA oil-immersion objective. FRAP analysis was carried out during S phase of cycle 11 or 12. We collected 0.5 μm thick confocal sections through the center of a selected centrosome. We bleached GFP signals using a focused 440 nm laser. We converted the Dendra2 signal using a focused 405 nm laser (targeted specifically at the central 4 pixels of the centrosome).