Figure 1. Detection of the phosphorylation of Arabidopsis thaliana MAPKs MPK6 and MPK3 upon treatment of seedlings for 15 min with the active epitope (flg22, 1 μM) of the bacterial elicitor flagellin (+). No phosphorylated MAPKs were detected in the control treated sample (-).

Materials and Reagents

1. Arabidopsis thaliana adult plants
   
2. Liquid nitrogen
   
3. Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Rabbit mAb #4370 (Cell Signaling Technology) (http://www.cellsignal.com/products/4370.html)
   
4. Anti-Rabbit secondary antibody
   
5. Tris-HCl
   
6. Glycerol
   
7. SDS
   
8. Dithiothreitol (DTT)
   
9. Bromphenol blue
   
10. Reagents for SDS-PAGE and Western Blotting
    
11. 6x Protein extraction/loading buffer (see Recipes)
    

Equipment

1. 1.5 ml Microcentrifuge tubes
   
2. Small petri dishes (~ 5 cm diameter) or 6-well plates
   
3. Cork borer (~ 10 mm diameter)
   
4. Glass beads (~ 1-2 mm diameter)
   
5. Silamat S6 (http://www.ivoclarvivadent.com/en/products/equipment/mixer/silamat-s6)
   
6. Alternatively for 4. and 5. Mortar (~ 5 cm diameter) and pestle
   
7. Vortexer
   
8. Heating block
   
9. Centrifuge
   
10. Equipment for SDS-PAGE, Western Blotting and detection
    

Procedure