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. 2021 Apr 8;72(12):4319–4332. doi: 10.1093/jxb/erab156

Fig. 4.

Fig. 4.

MYB21 recognizes and binds to the GARE elements of the FLS1 promoter. (A) Schematic diagrams of the FLS1 promoter constructs in yeast one-hybrid assays. Black triangles indicate GARE cis-elements (TTGTTA), and blue and orange triangles indicate the PY-BOX (TTTTTTCC) and TA-BOX (TATCCA), respectively. (B) Yeast one-hybrid assays of the interactions between MYB21 and FLS1 promoter fragments. The empty vectors of pGADT7-Rec were used as a negative control. (C) EMSA indicating that MYB21 binds to the GARE motifs. Recombinant GST-MYB21 was purified from E. coli. The promoter fragment containing the two GARE motifs of the FLS1 promoter was labeled with biotin. A competition assay for the protein–DNA binding was performed using 100× unlabeled wild-type (Competitor) and mutant probes. (D) ChIP enrichment of FLS1 promoter regions bound by MYB21-FLAG. Ten-day-old Pro35S:MYB21-FLAG and wild-type (Col-0) seedlings were used; DNA fragments were analyzed by quantitative PCR, with the Actin8 promoter as a reference. Enrichments in Pro35S:MYB21-FLAG were compared with wild-type seedlings. Error bars indicate the SD of three PCR repeats of four separate samples.