DEV02724 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. Expression pattern of Mixl1-GFP and Mixl1-lacZ. (A,B) Expression pattern of Mixl1-GFP and Mixl1-lacZ in early-streak, mid-streak and mid- to late-streak Mixl1^+/GFP and Mixl1^+/lacZ embryos. In the early-streak embryo, GFP and lacZ are expressed in the visceral endoderm, with a stronger lacZ expression in the posterior visceral endoderm. Both reporters are expressed (X-Gal staining being more visible than GFP) in the primitive streak and the nascent mesoderm of the mid-steak and mid- to late-streak embryos and in the visceral endoderm. (C) The GFP-expression pattern for staging of the gastrulation and guiding the harvest of cells for transplantation. (D) Transplanted cells in the anterior primitive streak of ARC/s host. (E) Visualization of cells that co-express GFP and lacZ after 24 hours of in vitro culture. (F,G) Presence of lacZ-expressing graft-derived Mixl1^+/GFP (F) and Mixl1^GFP/GFP (G) cells in the host embryo (left, ventral view; right, lateral view).

• Supplemental Figure 2 -

  Fig. S2. Effects on mesoderm and endoderm expansion in Mixl1 mutants. Expansion of the GFP-positive mesoderm during in vitro development of a Mixl1^+/GFP (A) and Mixl1^GFP/GFP (B) mutant embryo. The different fluorescence intensity correlates with the GFP gene dosage. Embryos were harvested at the mid-streak stage and cultured in vitro. The Mixl1^+/GFP embryo lost the GFP fluorescence after 12 hours of development, whereas the Mixl1^GFP/GFP embryo displayed strong fluorescence throughout the 24 hours of culture. (C) Mixl1^+/GFP and (D) Mixl1^GFP/GFP embryos with DiI-labelled endodermal cells visualized (as merged GFP and DiI fluorescent images) at 1 hour (left) and 24 hours (right) of in vitro culture.