Fig. S1

Expression analysis of nkx2.5 and Zs yellow in wild type and Tg(nkx2.5:Zs Yellow) animals.

(A,B) Lateral views of 72 hours post fertilization (hpf) wild type (A) or Tg(nkx2.5:Zs Yellow) (B) zebrafish embryos processed for in situ hybridization with nkx2.5 or ZsYellow riboprobes, respectively. Anterior to let. The brackets(*) highlight the locations of ventral and middle pharyngeal arch (PA) 1 and PA2 muscles as well as hypobranchial artery endothelium. The circles drawn with dotted lines show the location of dorsal PA1 and PA2 muscles. The arrows highlight the location fo the OFT, which is negative for nkx2.5(A) or ZsYellow (B) transcripts. The signals located dorsal to the heart in (B) are from cells associated with th pharyngeal arch arteries. (C-F) Ventral views of the heart and OFT in 72 hpf Tg(nkx2.5:ZsYellow) animals processed for in situ hybridization with a ZsYellow riboprobe (C) or double immunostained to detect ZsYellow (green) and Eln2+OFT smooth muscle cells (red; D-F). Anterior up. Merged (D) and single channel (E,F) confocal Z-stack are shown. The arrows highlight the ZsYellow+, Eln2+ OFT that protrudes from the ventricle, which is negative for ZsYellow transcripts at this stage(C). (G,H) Dorsal views of 28 hpf wild type (G) or Tg(nkx2.5:ZsYellow) (H) zebrafish embryos processed for in situ hybridization with nkx2.5 or ZsYellow riboprobes, respectively. The asterisk (*) highlights SHF progenitors for the distal ventricular myocardium. Little to no variability was observed in the greater than 30 animals examined. Numbers label the pharyngeal arches. Abbreviations: h, heart; l, liver; v, ventricle; a, atrium; oft, outflow tract. Scale Bars=50µm.