Fig. 3. Characterization of Ngn3-Nkx6.1 and Pdx1-Nkx6.1 transgene expression in the Nkx6.1 mutant background at E14.5. (A-D) Immunofluorescence detection of both Nkx6.1 (red; absent in Nkx6.1-null mice, B) and Ngn3 (green) shows strong co-expression in wild-type (A) and Nkx6.1^-/-;Ngn3-Nkx6.1 (C) pancreata, but weak or no co-expression in Nkx6.1^-/-;Pdx1-Nkx6.1 (D) pancreata. A subset of Ngn3⁺ cells is not targeted by the Ngn3-Nkx6.1 transgene (C). (E,F,H) Nkx6.1 (red; absent in Nkx6.1-null mice, F) is co-expressed with Pdx1 (green) in pancreata from wild-type (E) and Nkx6.1^-/-;Pdx1-Nkx6.1 (H) embryos. Notably, Pdx1⁺ acinar cells do not express Nkx6.1 protein in Nkx6.1^-/-;Pdx1-Nkx6.1 embryos (H, arrows). (G) Nkx6.1^-/-;Ngn3-Nkx6.1 embryos display a subset of Nkx6.1⁺ cells (red) with no expression of Pdx1 (green), and another subset with weak Pdx1 expression. (I,J,L) Nkx6.1 (red; absent in Nkx6.1-null mice, J) is co-expressed with insulin (Ins, green) in pancreata from wild-type (I) and Nkx6.1^-/-;Pdx1-Nkx6.1 (L, arrows) embryos. (K) Occasional co-localization of Nkx6.1 (red) and insulin (green) is also observed in Nkx6.1^-/-;Ngn3-Nkx6.1 pancreata (arrow). Scale bar: 50 µm.