Sequence and DNA-binding Activity of WRKY25. A. Amino acid sequence of WRKY25. The two WRKY motifs are indicated with the highly conserved WRKYGQK sequence and the residues forming the C2H2 zinc fingers underlined. B. Oligonucleotides used in the electrophoretic mobility shifting assay (EMSA). The Pchn5 probe contains two direct W-box repeats, while in the mPchn5 probe, the TTGACC sequences are mutated to TTGAAC. The wild-type and mutated W-box sequences are underlined. C. EMSA to test binding of recombinant WRKY25 to the W box motif in the Pchn5 probe. Binding reactions containing WRKY25 and Pchn5 produced two major DNA/protein complexes, which are indicated by arrows. Change of the TTGACC to TTGAAC in the mPchn5 probe abolished WRKY25 binding. No retarded bands were detected in the absence of the recombinant WRKY25 protein.