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Fig. 3. Morphological and functional analysis of Brg1^ep-/-(c) mutant fetuses. (A) Gross morphology of Brg1^L2/L2 (control; a) and Brg1^ep-/-(c) (b) E18.5 fetuses. fl, forelimb. (B) Hindlimb skeletal preparation from Brg1^L2/L2 (a) and Brg1^ep-/-(c) (b,c) E18.5 fetuses, stained for cartilage (Alcian Blue) and bone (Alizarin Red). Hindlimbs of two different mutants illustrate a mild (b) and the most severe (c) observed malformations. Two (b) and four (c) digits are missing in the autopods (au). The zeugopod (z) exhibits mild deformations of the tibia (ti) and fibula (fi) in b, and lacks the tibia in c. Note that the stylopod (st) and pelvic girdle (pg) are normal. fe, femur. (C) Whole-mount Fgf8 RNA in situ hybridization in control (a,c) and Brg1^ep-/-(c) (b,d) fetuses collected at E10.5 (a,b) and E11.5 (c,d). The arrowhead in b points to a mild abnormal domain of Fgf8 expression at E10.5. fl, forelimb; hl, hindlimb. (D) Anteroposterior transversal semi-thin sections of E11.5 (a) control (CT) and (b) Brg1^ep-/-(c) mutant hindlimb buds. The AER in control limb bud is indicated by an arrow. ^**, the altered AER region in the Brg1^ep-/-(c) mutant limb bud. Scale bar in b: 50 µm. (E) In vivo Lucifer yellow diffusion in (a) Brg1^L2/L2 and (b) Brg1^ep-/-(c) E18.5 fetuses. Dorsal skin sections were counterstained with DAPI (blue) and analysed by fluorescent microscopy. E, epidermis; D, dermis. Scale bar in a: 33 µm. (F) Barrier-dependent X-Gal diffusion assay on Brg1^L2/L2 (a) and Brg1^ep-/-(c) (b) E18.5 fetuses. (G) TEWL on dorsal and ventral skin of Brg1^L2/L2 and Brg1^ep-/-(c) E18.5 fetuses. ^***P<0.001.

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