Figure 5.

Overview of the protocol applied on Arabidopsis thaliana root protein extracts following treatment with 15 mM H₂O₂ for 30 min. (A) Immunoblot detection of MPK6 (bottom panel, arrow, MPK6), and phosphorylated MPK6 (top panel, top arrow, pMPK6) in conventional SDS-PAGE gels by means of anti-pTEpY antibody. In top panel of (A) the fainter band likely corresponds to phosphorylated MPK3 (bottom arrow, pMPK3) which is detectable depending on the separation efficiency of the gel. (B1,B2) Identical immunoblot from Phos-Tag™ separated samples in the presence of Zn²⁺ (B1) or Mn²⁺ (B2) immunodetected with antibody against MPK6 (B1,B2). Proteins were allowed to separate in the Phos-Tag™ gel for 4 h. (C1,C2) Identical immunoblots from Phos-Tag™ separated samples in the presence of Zn²⁺ (C1) or Mn²⁺ (C2). Proteins were allowed to separate for 6 h in the Phos-Tag™ gel. Panels on the left of (A) correspond to images of the protein ladder following Ponceau S staining of the membrane.