Zebrafish angiogenesis studies
To evaluate blood vessel formation in zebrafish, fertilized one-cell fli1a-EGFP transgenic lines embryos were injected with rapgef1a-MO and control-MO. At 48-hpf, embryos were dechorionated, anesthetized with 0.016% MS-222 (tricaine methanesulfonate, Sigma-Aldrich, St. Louis, MO). Zebrafish were then oriented on lateral side (anterior, left; posterior, right; dorsal, top), and mounted with 3% methylcellulose in a depression slide for observation by fluorescence microscopy. The phenotypes of complete intersegmental vessels (ISVs) (i.e. the number of ISVs that connect the DA to the DLAV) were quantitatively analyzed. The characterization of vasculature system and its development was described previously (Benedito et al., 2012; Covassin, Villefranc, Kacergis, Weinstein, & Lawson, 2006; Noguera-Troise et al., 2006; Ridgway et al., 2006; Siekmann & Lawson, 2007; Wythe et al., 2013).