Spinning disk confocal time-lapse imaging of the first mitosis in a C. elegans embryo expressing GFP-CeCENP-A in strain OD101. During prophase, discrete foci of GFP signal within the nucleus coalesce to form holocentric kinetochore plates. After nuclear envelope breakdown, chromosomes congress to the metaphase plate and are seen as paired lines. At metaphase, sister kinetochores form two lines facing opposite poles. The video is ∼12 min long, and the frame is ∼30 µm top to bottom. DIC is shown on the left, and GFP is shown on the right. Images were acquired at 10-s intervals.
Images were acquired via a spinning disk confocal microscope (CSU10; McBain Instruments) mounted on an inverted microscope (TE2000e; Nikon). Strain TH32 coexpressing GFP-histone H2b and GFP–γ-tubulin was imaged using a 60× 1.4 NA plan Apo objective with 1.5× auxiliary magnification and a cooled CCD camera (Orca ER; Hamamatsu) binning 2 × 2. Strain OD31 expressing GFP–KNL-2 was imaged in the same manner without the 1.5× auxiliary magnification.
|Spatial Axis||Image Size||Pixel Size|