Figure 2: 3D reconstruction of thread coiling within an immature GTC from the FIB-SEM data.

(a) Stacks of images from FIB-SEM were assembled into a 3D volume using Mimics software. This image was created by combining axial and transverse sections of the rendered volume with an SEM image of a separate complete thread skein. (b) Segmentation of a dozen continuous loops within a developing GTC revealed the precise 3D pattern of thread coiling (single loop in green). 3D reconstruction of the nucleus (light blue), nucleolus (dark blue) and mitochondria (maroon) shows the relative position of these structures to the developing thread. The loops shown are not the most recent ones laid down on the nucleus (these were too small to follow), but they do reflect nuclear shape at the time of synthesis. Inset shows the position of the rendered structures within the cell. (c) Based on the 3D structure of a single loop and its relationship to adjacent loops, our model (built in Maya 2013) reproduces the spiralling nature of the conical loops, the nesting of these conical structures, as well as the cabled appearance of the skein where the thread runs circumferentially along the skein surface.