2.8 ChIP-qPCR assay
Chromatin immunoprecipitation (ChIP)-qPCR assay was performed as described (Hong et al. , 2012; He et al. , 2021). Four-week-old leaves from plants expressing OsPHR2-Ov1 c fused with Flag-tag (Zhou et al ., 2008) were subjected to cross-linking for ChIP assays. Anti-Flag antibody was used for precipitation and negative IgG antibody as control. The precipitated cross-linked DNA was purified using EpiQuik™ plant ChIP kits. ChIP products were analyzed by RT-qPCR, and fold enrichment was calculated as the ratio of Flag-antibody/IgG–antibody quantifications. Error bars represent ±SE of three biological replicates. The OsACTIN2 gene promoter was used as a reference and the OsIPS1 gene promoter was set as positive control (Lv et al ., 2014). The primers used are listed in Table S1.