Figure 1—figure supplement 1

Figure 1—figure supplement 1

(A) Exemplary high-resolution fluorescence image of a blastoderm explant cross-section at bud stage. Interstitial fluid (grey) is marked by dextran cascade blue and mesendoderm tissue (green) by expression of gsc::GFP-CAAX. (B) Normalized distance of the main luminal cavity from the extension tip (a) or the explant back (b) (n = 90, N = 3). Schematic representation of the quantification method is shown at the top. (C) Cell diameter in stage-matched embryos and blastoderm explants from high to shield stage (high: embryo, number of cells = 270, n = 9, N = 3, explants, number of cells = 270, n = 9, N = 3; sphere: embryo, number of cells = 270, n = 9, N = 3, explants, number of cells = 270, n = 9, N = 3; 30% epiboly: embryo, number of cells = 240, n = 8, N = 3, explant, number of cells = 280, n = 9, N = 3; 50% epiboly: embryo, number of cells = 270, n = 9, N = 3, explant, number of cells = 270, n = 9, N = 3; germ ring: embryo, number of cells = 280, n = 9, N = 3, explant, number of cells = 270, n = 9, N = 3; shield: embryo, number of cells = 180, n = 6, N = 3, explant, number of cells = 240, n = 9, N = 3). ns, not significant (Kruskal-Wallis test). (D) Expression of two ZGA marker genes (chrd and tbx16) in stage-matched embryos (animal view) and blastoderm explants (side view) at sphere stage as determined by whole mount in situ hybridization. The proportion of embryos or blastoderm explants with a phenotype similar to the images shown is indicated in the lower right corner (chrd: embryos, n = 16, N = 3, explants, n = 33, N = 3; tbx16: embryos, n = 25, N = 3, explants, n = 45, N = 3). (E) High-resolution fluorescence images of krt4::EGFP-CAAX-expressing embryos (animal view) and blastoderm explants (side view) at bud stage. The proportion of embryos or blastoderm explants with a phenotype similar to the images shown is indicated in the lower right corner (embryos: n = 8, N = 2; explants: n = 9, N = 2). (F) Intensity of krt4::EGFP-CAAX in stage-matched embryos and blastoderm explants from late sphere stage onwards (embryos: n = 7, N = 3; explants: n = 8, N = 3). Time in min. (G) Expression of additional mesendoderm (gsc, flh, tbx6 and myoD) and endoderm (sox32) marker genes as determined by whole mount in situ hybridization of bud stage embryos and blastoderm explants. Schematic representation of the different views for embryos and blastoderm explants is shown on the left. The proportion of embryos or blastoderm explants with a phenotype similar to the images shown is indicated in the lower right corner (gsc: embryos, n = 46, N = 4, explants, n = 58, N = 6; flh: embryos, n = 17, N = 2, explants, n = 21, N = 2; tbx6: embryos, n = 27, N = 3, explants, n = 22, N = 3; myoD: embryos, n = 20, N = 3, explants, n = 20, N = 3; sox32: embryos, n = 22, N = 3, explants, n = 18, N = 3). (H) Percentage of bud stage blastoderm explants expressing or not neuroectoderm (six3: n = 26, N = 4), ectoderm (gata2: n = 33, N = 6), mesendoderm (hgg: n = 49, N = 5; gsc: n = 58, N = 6; myoD: n = 20, N = 3; ntl: n = 48, N = 4; flh: n = 21, N = 2; papc: n = 55, N = 5; tbx6: n = 22, N = 3) and endoderm (sox32: n = 18, N = 3) marker genes as determined by whole mount in situ hybridization. (I) Normalized expression domain of additional mesendoderm marker genes (gsc: n = 22, N = 6; flh: n = 17, N = 2; tbx6: n = 17, N = 3; myoD: n = 7, N = 3) along the back-tip axis of bud stage blastoderm explants. (J) Percentage of blastoderm explants showing expression of sox32 in a single cell cluster, a cluster and some more individual cells, only individual cells or no expression at bud stage as determined by whole mount in situ hybridization (n = 18, N = 3). White asterisks denote the main luminal cavity in explants. Scale bars: 100 µm (A, E), 200 µm (D, G).

Figure 1—figure supplement 1—source data 1.