a, Plot of secGFP pool fluorescence intensity upon micropuncture (green). Kymographs show the time-lapse imaging of the secGFP pool used for the plot. b, c, Luminal Fgf3–GFP signal recovery of whole pool bleached (left) and micro-punctured (right) organs during 48 min of acquisition. Kymographs show time-lapse imaging of Fgf3–GFP pool. Single time points of time-lapse imaging after micro-puncture (right). Scale bar, 5 μm. d, Quantification of pea3 transcript levels at t = 0 h, 1 h and 4 h after micropuncture of organ 2 expressing lexOP:fgf3–GFP. Unperturbed organ 3 was used for normalization. Comparison of control and punctured organs suggests that pea3 levels are normal immediately after puncture, are reduced 1 h later and recovered by 4 h (N0 h puncture = 6, N0 h control = 5, N1 h puncture = 6, N1 h control = 5, N4 h puncture = 6, N4 h control = 5, P0 h = 0.7922, P1 h = 0.0043, P4 h = 0.4286). e, Organ deposition delay upon lumina micropuncture of secGFP-expressing second and third organs (Nctrl second organ = 22, Npuncture second organ = 23, Nctrl third organ = 8, Npuncture third organ = 9, Psecond organ = 6.928 × 10−6, Pthird organ = 0.0061). Scale bar, 200 µm. f, g, Shroom3 morphant primordia show intervals with no or delayed deposition. f, Kymographs of shroom3 MO and control. Scale bars, 200 µm, 5 h. g, Organ pattern in shroom3 MO and control at 2 d.p.f. Scale bar, 200 µm.