For Mtz treatment, the Tg(lfabp:DenNTR) transgenic larvae at 5 dpf were incubated with 10 mM Mtz (metronidazole, Sigma) for 24 hours. For ethanol (EtOH) treatment, larvae were treated with 1.5% EtOH (Sigma Aldrich) for 48 hours, starting from 4 dpf17 (link). For HU (hydroxyurea, Sigma) treatment, embryos were treated with 100 mM HU in egg water from R0h to R8h. Larvae were treated with 0.1 μM Chk1i (CHIR-124, Selleck), 5 μM ATRi (VE-821, Selleck), 2 μM ATMi (KU-55933, Selleck) and 20 μM Mirin (Selleck) from R8h to R48h. The chemical solutions were renewed every 24 h to maintain the pharmacological effects. And a 0.2% DMSO solution in egg water was used as a control. When chemical treatment is removed, the larvae need to be washed and recovered in the egg water.
For temporal control of CreER activities, larvae were collected and incubated with 5 μM 4-hydroxytamoxifen (4-OHT, Sigma) for 24 h from 4 to 5 dpf, followed by three washes with egg water.
The larvae of Tg(hsp70l:nbn-p2A-mCherry)cq141 were heat-shocked at 38.5 °C for 40 min at the indicated time frame and returned to 28.5 °C for further analysis.
For BODIPY feeding assay, larvae were fed with BODIPY FL C5 (Invitrogen) for 4 h at 5 dpf and then live imaging, as previously described26 (link).
For temporal control of CreER activities, larvae were collected and incubated with 5 μM 4-hydroxytamoxifen (4-OHT, Sigma) for 24 h from 4 to 5 dpf, followed by three washes with egg water.
The larvae of Tg(hsp70l:nbn-p2A-mCherry)cq141 were heat-shocked at 38.5 °C for 40 min at the indicated time frame and returned to 28.5 °C for further analysis.
For BODIPY feeding assay, larvae were fed with BODIPY FL C5 (Invitrogen) for 4 h at 5 dpf and then live imaging, as previously described26 (link).
Free full text: Click here
