EMSA

EMSA assay was based on LightShift Chemiluminescent EMSA Kit (Thermo Fisher Scientific, 20148) with some modifications (Jiang et al. 2022). One μg purified recombinant protein His-ICE1 was incubated with biotin-labeled probes at 25 °C for 30 min. The binding reactions were terminated by the addition of 2 μL 10× loading buffer [250 mM Tris-HCl, pH 8.0, and 40% (v/v) glycerol], which were then separated on 8% native polyacrylamide gels in 0.5×TBE buffer. The signals were detected with HRP-conjugated anti-Biotin antibody (Thermo Fisher 89880, 1:10,000 dilution). The primers used in this assay were shown in Supplemental Table S1.