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Fig. 1. Noonan-associated mutation of SHP-2 leads to heart defects in Xenopus. (A) Schematic of SHP-2 constructs. Note that constructs used in Xenopus in vivo experiments were not epitope tagged with HA. (B) Western blot analysis of embryo lysates from embryos injected with the respective HA epitope-tagged SHP-2 constructs at stages 10 (gastrula), 16 (early neurula) and 26 (late neurula) probed with an anti-HA antibody, an anti-SHP-2 antibody that recognizes both endogenous and introduced versions of SHP-2, and an anti-EF-2 antibody as a loading control. (C-H) Whole-mount antibody staining of cleared stage 36 embryos with anti-tropomyosin antibody (Tmy). Anterior is to the left, posterior to the right. Scale bar: 100 μm. (I-Q) Whole-mount in situ hybridization of cleared stage 33 from uninjected (I,L,O), SHP-2FL (2 ng; J,M,P) and SHP-2N308D (2 ng; K,N,Q) embryos probed for Nkx2.5 (I-K), Tbx5 (L-N) or Tbx20 (O-Q). Scale bar: 1 mm. a, atrium; oft, outflow tract; v, ventricle. |
