Xenopus Embryo gpx3 Knockdown

The gpx3 MO was injected into the Xenopus embryos ventral region at the 4-cell stage. The MO-injected embryos were collected at the desired stage and were fixed in fixative MEMFA (4% paraformaldehyde, 0.1 M MOPS buffer (pH 7.4), 1 mM MgSO₄, 2 Mm ethylene glycol-bis(β-amino ethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA) overnight at 4 °C. The embryos were dehydrated before storage in 100% methanol at −20 °C. To prepare the antisense digoxigenin (DIG)-labeling probes, DNA templates were linearized using appropriate restriction enzymes. Probes were synthesized using SP6 or T7 RNA polymerase (Ambion) and were detected using an alkaline phosphatase-labeled antidigoxigenin antibody (1:1000, Roche, Basel, Switzerland) or nitro blue tetrazolium/5-bromo-4-chloro-3indolyl phosphate (NBT/BCIP) [25 (link)].