Figure 3

Stages 4-6 of C. salei. All scale bars 200 μm. Sytox staining, a-d, f-g; light micrograph, e. a-b, Stage 4, Primary thickening. a: The blastopore region (evident in Figures 2h-j) consists of several dense layers of nuclei. It bulges slightly outwards, a phenomenon which in this and subsequent figures is called primary thickening (PT). The surrounding nuclei are arranged in irregular patches (arrows). b: Same egg as in a, seen from the opposite side. At this pole of the egg, the nuclei are more evenly distributed. c-e, Stage 5, Cumulus migrating. c: From the primary thickening, a large cell cluster (cumulus, Cu) starts to separate from the remaining rest of the primary thickening (PT). d: Due to the ongoing migration (white arrow) of the cumulus (Cu), the distance to the primary thickening (PT) increases. e: Living egg (egg number 1 from Additional file 2 at movie frame 960) in late stage 5. The cumulus (Cu) has reached its final position after separation from the primary thickening (PT). The migration of cells from the primary thickening beneath the surface layer has resulted in one hemisphere with greater cell density (top of photo e) compared with the other hemisphere (bottom of photo e). The line of demarcation between the dense hemisphere and the less dense one is the 'equator'. This marked difference in cell density is evident between the two arrowheads in e. f-h, Stage 6, Dorsal field. f: After the cumulus has reached the equator, it disintegrates. The tissue between the cumulus and the primary thickening spreads laterally, forming a region of low cell density called the dorsal field (DF). This region will continue to have much yolk in further stages of development. g: Same egg as in f rotated 120 degrees. The primary thickening (PT) is at the posterior edge of the developing embryo body and located in the hemisphere with greater cell density. The hemisphere with lower cell density is extra-embryonic (Ee) since it continues as a yolk-filled region. h: Posterior view of the primary thickening (PT). The dorsal field (DF) expands to about 100 degrees in width, and the cumulus has disappeared.