Series 700 halocarbon oil

Aligned embryos on coverslips were covered with Series 700 halocarbon oil (Sigma-Aldrich). Imaging of embryos after mRNA injection (Figure 1c) was performed with a spinning disc Revolution XD microscope (Andor, UK) at 22°C. Stacks of around 20 frames 1 μm were taken at indicated times using a 100 × 1.4 oil immersion objective (Nikon, Japan) and iXon +512 EMCCD camera (Andor). Time-lapse microscopy was performed with an inverted wide-field DeltaVision microscope (Applied Precision Inc., Issaquah, WA) at 18–20°C in a temperature-controlled room. One stack of ~20 frames (0.8 μm apart) was acquired every 1 or 2 min using a 100 × 1.4 oil immersion objective (Olympus, Japan) and an EMCCD camera (Roper Cascade 1024, Roper Technologies, Inc., Sarasota, FL). Widefield images were restored by deconvolution with the Huygens v15.10/16.10 deconvolution software using a calculated point-spread function (RRID:SCR_014237, Scientific Volume Imaging, The Netherlands). Movies were assembled using FIJI software (RRID:SCR_002285) (Schindelin et al., 2012 (link)) and selected stills were processed with Photoshop CS6 (Adobe Systems Incorporated, San Jose, CA).

Microinjection experiments were performed as previously described (Oliveira et al., 2010 (link); Piskadlo et al., 2017 (link)). Dechorionated embryos (1–1.5 h old) were glued to a #1.5 coverslip and covered with Series 700 halocarbon oil (H8898; Sigma-Aldrich). Embryos were then injected at 18–20°C into the posterior pole using a Burleigh Thorlabs micromanipulator, a Femtojet microinjection system (Eppendorf), and prepulled Femtotip I needles (Eppendorf). Injections were performed using 12 mg/ml UbcH10^C114S diluted in 20 mM Tris-HCl, pH 7.5, 300 mM NaCl, 6 mg/ml TEV protease in TEV buffer (20 mM Tris-HCl, pH 8.0, 1 mM EDTA, 50 mM NaCl, and 2 mM DTT), or 2 mM colchicine diluted in PBS, pH 7.4. TALE-light GFP 1.686 was produced as previously described (Yuan et al., 2014 (link)) and was injected at 1 mg/ml in 40 mM Hepes and 150 mM KCl, pH 7.4. Aurora B–EGFP mRNA preparation/injection was performed as previously described (Oliveira et al., 2010 (link)) and used at a concentration of 74 ng/µl.