Fig. 1 Basic principle of EMSA experiment Asterisks indicate labeled probes，TFs indicate transcription factors，and TFBS indicate transcription factor binding sites

Fig. 2 Basic principle of DNase I footprint technology Adding appropriate concentration of deoxyribonuclease I（DNase I）to label strand in the DNA fragment，the DNA motif bound to the target protein will not be hydrolyzed by DNase I. After comparing the autoradiograph of the DNA fragment not bound to the target protein，interrupted DNA gradient bands can be obtained

Fig. 3 Basic principle of ChIP experiment Cross-link the target protein with DNA in vivo through a cross-linking agent，and then fragment the DNA and protein complex. After that，precipitate with the specific antibody of the target protein to enrich the DNA fragments bound to the target protein. Reversal of crosslink and DNA extraction，finally analysis of purified DNA

Fig. 4 Basic principle of MST experiment A：Fluorescence detector. B：MST optical element schematic diagram. C：MST experiment signal curve. D：MST combination experiment

Fig. 5 Basic principle of ITC experiment The titration of reactants of known concentration into the sample causes the reaction between the components in the sample cell to be endothermic or exothermic. The temperature compensation system maintains a constant temperature difference between the sample cell and the reference cell，and the data are recorded to simulate the integrated heating isotherm. After that，information about the reaction enthalpy change，binding affinity and binding stoichiometry can be obtained.

Fig. 6 Basic principle of bacterial one hybrid system A：Transcription factor recognition binding sites activate the expressions of the reporter genes. B：Construct α-TF expression vector and binding site library respectively. C：Eliminate self-activation to purify binding site library. D：Selection and analysis of binding sites