Fig. 4.

miR173 triggers phased siRNA formation from modified GFP transcripts. (A) Organization of the GFP-based constructs with engineered miRNA target sites. The proportion of cloned 5′ RACE products corresponding to cleavage at a site is shown above the arrows. Black arrows indicate the canonical miRNA-guided cleavage position. (B) Blot assays. TAS1c- and GFP-based constructs containing miR173 or miR171 target sites were expressed or coexpressed as indicated above the blot panels. One of two biological replicates is shown. EtBr-stained 5S rRNA/tRNA and 18S/25S rRNA are shown as loading controls. (C) Flow chart. (D and E) Twenty-one-nucleotide small RNA read distributions across 173-GFP and HYG^R transcripts. (Insets) Radar plots display percentages of reads corresponding to each of the 21 registers, with the 5′ end formed by miR173-guided cleavage or the transcription start site in 173-GFP and HYG^R, respectively, defined as register 1.