Dullard^−/− embryos fail to form primordial germ cells (PGCs).

(A–H) Absence of alkaline phosphatase (AP)-positive PGCs in Dullard^−/− embryos at the (A, B) E7.75 neural plate stage (A′, B′; higher magnification views) and (C–D) E8.5 early somite stage (C′, D′; higher magnification views). (E–H) Histology of E7.5 (E, E′) Dullard ^+/− and (F, F′) Dullard^−/− embryos revealing the absence of AP-positive cells in the endoderm of Dullard^−/− embryos, while few positive cells in the mesoderm displayed (H) the characteristic staining pattern (in the cell membrane and a cytoplasmic aggregate) of PGCs in the endoderm of (G) Dullard ^+/− embryos. (I, J) Markedly reduced population of Dppa3-expressing cells in the E7.5 Dullard^−/− embryo (confocal images). (K, L′′) No clustering of Ifitm3-positive cells in the E7.5 Dullard^−/− embryo. (K, L; whole embryo bright-field view; K′, L′; immunostaining, K′′, L′′; high magnification views of Iftim3-positive (red) cells in the boxed areas of K′ and L′; DAPI nuclear staining (blue). (M–N) Marked reduction in the numbers of Prdm1-positive cells in the E7.25 streak-stage Dullard^−/− embryo (M′, N′; high magnification views of the boxed areas in M, N, respectively). (O–P′′) Down-regulation of Ifitm1 expression in the posterior mesoderm of the E7.5 Dullard^−/− embryo [O, P, whole embryo; O′, P′, merged bright-field and immunofluorescence images (boxed area); O′′, P′′, immunofluorescence image only]. (A, A′, C, C′, E, E′, G, I, K–K′′, M, M′, O–O′′) Dullard ^+/− embryos; (B, B′, D, D′, F, F′, H, J, L–L′′, N, N′, P–P′′) Dullard^−/− embryos. (E, F) Dashed lines indicate the plane of sectioning. (E′, F′) Dashed lines mark the basal surface of the endoderm layer. Scale bars = 100 µm (A–F′, K–L′, M–P′′), 50 µm (I, J) and 25 µm (K′′, L′′).