Zebrafish and their embryos were handled according to standard protocols23 and in accordance with University of Massachusetts Medical School IACUC guidelines. For laser-assisted microsurgery, embryos at 46 hpf were anesthetized and immobilized in 0.5% of low-melt agarose (Biorad). The connection between AA5 and AA6 and the ventral aorta was ablated using a Micropoint laser (Photonic Instrument, Inc) mounted on a Zeiss AX10 Imager M1. SU5416 (Calbiochem) was prepared and used as described previously11 (link). Control embryos were treated with 0.1% dimethyl sulfoxide (DMSO). To arrest heartbeat, embryos were treated with 15 mM of 2,3-butanedione 2-monoxime (BDM; Sigma-Aldrich) or with buffered Tricaine methanesulfonate (Sigma-Aldrich) at 0.66 mg/ml in egg water for the indicated times. Two-photon time-lapse imaging, confocal microscopy and microangiography was performed as previously13 (link), 24 (link), with additional modifications as noted in Supplementary Methods. Antisense riboprobes against dll4, vegfa, kdrl, fli1a, and cdh5 were generated and used for whole mount in situ hybridization as described elsewhere25 (link). A klf2a fragment was PCR amplified and cloned by Gateway recombination. The resulting clone was linearized with BglII and a DIG-labeled riboprobe was synthesized using T7 polymerase. Digoxigenin (DIG)-labeled locked nucleic acid (LNA) probes (Exiqon, Copenhagen) were used to detect mature miR-126 and let-7 using in situ hybridization or Northern analysis as described elsewhere18 (link). Morpholinos, mRNA and Tol2-based plasmids were prepared and injected as previously11 (link),21 (link). In cases of co-injection with Morpholinos, Tol2-plasmids and transposase, a DNA/transposase mRNA mixture was initially injected, followed by Morpholino. Plasmid construction details are provided in the full methods section. Morpholinos against vegfa, tnnt2 and gata1 have been described elsewhere15 (link), 26 (link), 25 (link); all other Morpholino and oligonucleotide sequences are provided in the full methods section.