Fig. 6

OsHsfB4d directly binds to the promoter of OsHsp18.0-CI. a Diagram of the positions of the primers and probes used in the EMSA and ChIP-qPCR experiments. b EMSA using recombinant His-tagged OsHsfB4d and the various deletion derivatives of the OsHsp18.0-CI promoter fragment. Lane 1 contained only the free probe. Lanes 2–7 contained the purified His-tagged OsHsfB4d and the probe. For the competitive EMSA, the purified OsHsfB4d protein was preincubated with 5×, 10×, 50×, 200× molar excess of unlabeled probe 1 (lane 3–6), 5× probe 2 (lane 8) and 5× probe 3 fragments (lane 10) before the addition of the corresponding biotin-labelled promoter fragment. c Binding of OsHsfB4d to the perfect HSE cis-element-containing fragment in a ChIP-qPCR assay. ChIP and EMSA experiment used the same primer pairs. The Actin was used as a nonspecific target gene