Microinjection of Drosophila Embryos

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.