Fig. S2

Further characterization and validation of two independent morpholinos targeting tgfβR2 and analysis of primitive haematopoiesis in tgfβR2 morphants. (A) Schematic representation of the genomic organization of the tgfbR2 gene, location of morpholinos and primers used to validate the morpholinos. (B) Western blot against TgfβR2 showing a decrease of WT protein induced by the tgfbR2^MO1 morpholino. (C) Validation of the tgfbR2^MO2 by qPCR on 24hpf cDNA with tgfbR2 F2/R2 primers (see below). Ef1a PCR was used as a control for the PCR. (D) Expression of runx1, cmyb and ikzf1 at 48hpf is reduced in the trunk and CHT of tgfbR2 morphants. Expression of cmyb, ikzf1 and lplastin is severely reduced (E) in the thymus and (F) in the CHT of tgfbR2 morphants at 4dpf. (G) Characterisation of the tgfbR2 MO1 and MO2 morpholinos. Runx1 expression is decreased upon injection of either morpholino, whereas the arterial markers dll4, notch3 and dlC and the vascular markers kdrl and fli1 are grossly normal. (H) Expression of scl, gata1 and pu.1 is indistinguishable between wildtype and tgfbR2 morphant embryos at 20hpf. (I) Expression of the primitive hematopoietic markers gata1 and pu.1 is unaffected in tgfbR2 morphants at 24hpf. (J) tgfbR2 morphants show a slight reduction in odianisidine staining at 36hpf. Numbers of embryos analysed are shown in each panel as number of affected embryos/total observed.